This pathway, SARS-CoV-2 infection of human cells (COVID-19), was initially generated via electronic inference from the manually curated and reviewed Reactome SARS-CoV-1 (Human SARS coronavirus) infection pathway. The inference process created SARS-CoV-2 events corresponding to each event in the SARS-CoV-1 pathway and populated those events with SARS-CoV-2 protein-containing physical entities based on orthology to SARS-CoV-1 proteins (https://reactome.org/documentation/inferred-events). All of these computationally created events and entities have been reviewed by Reactome curators and modified as appropriate where recently published experimental data indicate the existences of differences between the molecular details of the SARS-CoV-1 and SARS-CoV-2 infection pathways.
SARS‑CoV‑2 infection begins with the binding of viral S (spike) protein to cell surface angiotensin converting enzyme 2 (ACE2) and endocytosis of the bound virion. Within the endocytic vesicle, host proteases mediate cleavage of S protein into S1 and S2 fragments, leading to S2‑mediated fusion of the viral and host endosome membranes and release of the viral capsid into the host cell cytosol. The capsid is uncoated to free the viral genomic RNA, whose cap‑dependent translation produces polyprotein pp1a and, by means of a 1‑base frameshift, polyprotein pp1ab. Autoproteolytic cleavage of pp1a and pp1ab generates 15 or 16 nonstructural proteins (nsps) with various functions. Importantly, the RNA dependent RNA polymerase (RdRP) activity is encoded in nsp12. Nsp3, 4, and 6 induce rearrangement of the cellular endoplasmic reticulum membrane to form cytosolic double membrane vesicles (DMVs) where the viral replication transcription complex is assembled and anchored. With viral genomic RNA as a template, viral replicase‑transcriptase synthesizes a full length negative sense antigenome, which in turn serves as a template for the synthesis of new genomic RNA. The replicase‑transcriptase can also switch template during discontinuous transcription of the genome at transcription regulated sequences to produce a nested set of negative‑sense subgenomic (sg) RNAs, which are used as templates for the synthesis of positive‑sense sgRNAs that are translated to generate viral proteins. Finally, viral particle assembly occurs in the ER Golgi intermediate compartment (ERGIC). Viral M protein provides the scaffold for virion morphogenesis (Hartenian et al. 2020; Fung & Liu 2019; Masters 2006).
View original pathway at Reactome.
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Muramatsu T, Takemoto C, Kim YT, Wang H, Nishii W, Terada T, Shirouzu M, Yokoyama S.; ''SARS-CoV 3CL protease cleaves its C-terminal autoprocessing site by novel subsite cooperativity.''; PubMedEurope PMCScholia
Bhardwaj K, Sun J, Holzenburg A, Guarino LA, Kao CC.; ''RNA recognition and cleavage by the SARS coronavirus endoribonuclease.''; PubMedEurope PMCScholia
Zhou N, Pan T, Zhang J, Li Q, Zhang X, Bai C, Huang F, Peng T, Zhang J, Liu C, Tao L, Zhang H.; ''Glycopeptide Antibiotics Potently Inhibit Cathepsin L in the Late Endosome/Lysosome and Block the Entry of Ebola Virus, Middle East Respiratory Syndrome Coronavirus (MERS-CoV), and Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV).''; PubMedEurope PMCScholia
Ujike M, Huang C, Shirato K, Matsuyama S, Makino S, Taguchi F.; ''Two palmitylated cysteine residues of the severe acute respiratory syndrome coronavirus spike (S) protein are critical for S incorporation into virus-like particles, but not for M-S co-localization.''; PubMedEurope PMCScholia
de Haan CA, Kuo L, Masters PS, Vennema H, Rottier PJ.; ''Coronavirus particle assembly: primary structure requirements of the membrane protein.''; PubMedEurope PMCScholia
Hatakeyama S, Matsuoka Y, Ueshiba H, Komatsu N, Itoh K, Shichijo S, Kanai T, Fukushi M, Ishida I, Kirikae T, Sasazuki T, Miyoshi-Akiyama T.; ''Dissection and identification of regions required to form pseudoparticles by the interaction between the nucleocapsid (N) and membrane (M) proteins of SARS coronavirus.''; PubMedEurope PMCScholia
Zhou D, Tian X, Qi R, Peng C, Zhang W.; ''Identification of 22 N-glycosites on spike glycoprotein of SARS-CoV-2 and accessible surface glycopeptide motifs: implications for vaccination and antibody therapeutics.''; PubMedEurope PMCScholia
Baranov PV, Henderson CM, Anderson CB, Gesteland RF, Atkins JF, Howard MT.; ''Programmed ribosomal frameshifting in decoding the SARS-CoV genome.''; PubMedEurope PMCScholia
Ritchie G, Harvey DJ, Feldmann F, Stroeher U, Feldmann H, Royle L, Dwek RA, Rudd PM.; ''Identification of N-linked carbohydrates from severe acute respiratory syndrome (SARS) spike glycoprotein.''; PubMedEurope PMCScholia
Siu YL, Teoh KT, Lo J, Chan CM, Kien F, Escriou N, Tsao SW, Nicholls JM, Altmeyer R, Peiris JS, Bruzzone R, Nal B.; ''The M, E, and N structural proteins of the severe acute respiratory syndrome coronavirus are required for efficient assembly, trafficking, and release of virus-like particles.''; PubMedEurope PMCScholia
Sakai Y, Kawachi K, Terada Y, Omori H, Matsuura Y, Kamitani W.; ''Two-amino acids change in the nsp4 of SARS coronavirus abolishes viral replication.''; PubMedEurope PMCScholia
Tohge T, Nishiyama Y, Hirai MY, Yano M, Nakajima J, Awazuhara M, Inoue E, Takahashi H, Goodenowe DB, Kitayama M, Noji M, Yamazaki M, Saito K.; ''Functional genomics by integrated analysis of metabolome and transcriptome of Arabidopsis plants over-expressing an MYB transcription factor.''; PubMedEurope PMCScholia
Heurich A, Hofmann-Winkler H, Gierer S, Liepold T, Jahn O, Pöhlmann S.; ''TMPRSS2 and ADAM17 cleave ACE2 differentially and only proteolysis by TMPRSS2 augments entry driven by the severe acute respiratory syndrome coronavirus spike protein.''; PubMedEurope PMCScholia
Li W, Moore MJ, Vasilieva N, Sui J, Wong SK, Berne MA, Somasundaran M, Sullivan JL, Luzuriaga K, Greenough TC, Choe H, Farzan M.; ''Angiotensin-converting enzyme 2 is a functional receptor for the SARS coronavirus.''; PubMedEurope PMCScholia
Wu F, Zhao S, Yu B, Chen YM, Wang W, Song ZG, Hu Y, Tao ZW, Tian JH, Pei YY, Yuan ML, Zhang YL, Dai FH, Liu Y, Wang QM, Zheng JJ, Xu L, Holmes EC, Zhang YZ.; ''A new coronavirus associated with human respiratory disease in China.''; PubMedEurope PMCScholia
Surjit M, Kumar R, Mishra RN, Reddy MK, Chow VT, Lal SK.; ''The severe acute respiratory syndrome coronavirus nucleocapsid protein is phosphorylated and localizes in the cytoplasm by 14-3-3-mediated translocation.''; PubMedEurope PMCScholia
Leduc R, Molloy SS, Thorne BA, Thomas G.; ''Activation of human furin precursor processing endoprotease occurs by an intramolecular autoproteolytic cleavage.''; PubMedEurope PMCScholia
Al-Bari MA.; ''Chloroquine analogues in drug discovery: new directions of uses, mechanisms of actions and toxic manifestations from malaria to multifarious diseases.''; PubMedEurope PMCScholia
Takeda M, Chang CK, Ikeya T, Güntert P, Chang YH, Hsu YL, Huang TH, Kainosho M.; ''Solution structure of the c-terminal dimerization domain of SARS coronavirus nucleocapsid protein solved by the SAIL-NMR method.''; PubMedEurope PMCScholia
He R, Leeson A, Ballantine M, Andonov A, Baker L, Dobie F, Li Y, Bastien N, Feldmann H, Strocher U, Theriault S, Cutts T, Cao J, Booth TF, Plummer FA, Tyler S, Li X.; ''Characterization of protein-protein interactions between the nucleocapsid protein and membrane protein of the SARS coronavirus.''; PubMedEurope PMCScholia
Taylor DL, Kang MS, Brennan TM, Bridges CG, Sunkara PS, Tyms AS.; ''Inhibition of alpha-glucosidase I of the glycoprotein-processing enzymes by 6-O-butanoyl castanospermine (MDL 28,574) and its consequences in human immunodeficiency virus-infected T cells.''; PubMedEurope PMCScholia
Yin W, Mao C, Luan X, Shen DD, Shen Q, Su H, Wang X, Zhou F, Zhao W, Gao M, Chang S, Xie YC, Tian G, Jiang HW, Tao SC, Shen J, Jiang Y, Jiang H, Xu Y, Zhang S, Zhang Y, Xu HE.; ''Structural basis for inhibition of the RNA-dependent RNA polymerase from SARS-CoV-2 by remdesivir.''; PubMedEurope PMCScholia
Li FQ, Xiao H, Tam JP, Liu DX.; ''Sumoylation of the nucleocapsid protein of severe acute respiratory syndrome coronavirus.''; PubMedEurope PMCScholia
Belouzard S, Chu VC, Whittaker GR.; ''Activation of the SARS coronavirus spike protein via sequential proteolytic cleavage at two distinct sites.''; PubMedEurope PMCScholia
Glowacka I, Bertram S, Müller MA, Allen P, Soilleux E, Pfefferle S, Steffen I, Tsegaye TS, He Y, Gnirss K, Niemeyer D, Schneider H, Drosten C, Pöhlmann S.; ''Evidence that TMPRSS2 activates the severe acute respiratory syndrome coronavirus spike protein for membrane fusion and reduces viral control by the humoral immune response.''; PubMedEurope PMCScholia
Locker JK, Opstelten DJ, Ericsson M, Horzinek MC, Rottier PJ.; ''Oligomerization of a trans-Golgi/trans-Golgi network retained protein occurs in the Golgi complex and may be part of its retention.''; PubMedEurope PMCScholia
Watanabe Y, Allen JD, Wrapp D, McLellan JS, Crispin M.; ''Site-specific glycan analysis of the SARS-CoV-2 spike.''; PubMedEurope PMCScholia
Saikatendu KS, Joseph JS, Subramanian V, Neuman BW, Buchmeier MJ, Stevens RC, Kuhn P.; ''Ribonucleocapsid formation of severe acute respiratory syndrome coronavirus through molecular action of the N-terminal domain of N protein.''; PubMedEurope PMCScholia
Chang CK, Hsu YL, Chang YH, Chao FA, Wu MC, Huang YS, Hu CK, Huang TH.; ''Multiple nucleic acid binding sites and intrinsic disorder of severe acute respiratory syndrome coronavirus nucleocapsid protein: implications for ribonucleocapsid protein packaging.''; PubMedEurope PMCScholia
Cohen JR, Lin LD, Machamer CE.; ''Identification of a Golgi complex-targeting signal in the cytoplasmic tail of the severe acute respiratory syndrome coronavirus envelope protein.''; PubMedEurope PMCScholia
Amirian ES, Levy JK.; ''Current knowledge about the antivirals remdesivir (GS-5734) and GS-441524 as therapeutic options for coronaviruses.''; PubMedEurope PMCScholia
Ujike M, Taguchi F.; ''Incorporation of spike and membrane glycoproteins into coronavirus virions.''; PubMedEurope PMCScholia
Li F, Li W, Farzan M, Harrison SC.; ''Structure of SARS coronavirus spike receptor-binding domain complexed with receptor.''; PubMedEurope PMCScholia
Ravindra NG, Alfajaro MM, Gasque V, Wei J, Filler RB, Huston NC, Wan H, Szigeti-Buck K, Wang B, Montgomery RR, Eisenbarth SC, Williams A, Pyle AM, Iwasaki A, Horvath TL, Foxman EF, van Dijk D, Wilen CB.; ''Single-cell longitudinal analysis of SARS-CoV-2 infection in human bronchial epithelial cells.''; PubMedEurope PMCScholia
Oostra M, te Lintelo EG, Deijs M, Verheije MH, Rottier PJ, de Haan CA.; ''Localization and membrane topology of coronavirus nonstructural protein 4: involvement of the early secretory pathway in replication.''; PubMedEurope PMCScholia
Yu X, Chen S, Hou P, Wang M, Chen Y, Guo D.; ''VHL negatively regulates SARS coronavirus replication by modulating nsp16 ubiquitination and stability.''; PubMedEurope PMCScholia
Toots M, Yoon JJ, Cox RM, Hart M, Sticher ZM, Makhsous N, Plesker R, Barrena AH, Reddy PG, Mitchell DG, Shean RC, Bluemling GR, Kolykhalov AA, Greninger AL, Natchus MG, Painter GR, Plemper RK.; ''Characterization of orally efficacious influenza drug with high resistance barrier in ferrets and human airway epithelia.''; PubMedEurope PMCScholia
Voss D, Kern A, Traggiai E, Eickmann M, Stadler K, Lanzavecchia A, Becker S.; ''Characterization of severe acute respiratory syndrome coronavirus membrane protein.''; PubMedEurope PMCScholia
Marra MA, Jones SJ, Astell CR, Holt RA, Brooks-Wilson A, Butterfield YS, Khattra J, Asano JK, Barber SA, Chan SY, Cloutier A, Coughlin SM, Freeman D, Girn N, Griffith OL, Leach SR, Mayo M, McDonald H, Montgomery SB, Pandoh PK, Petrescu AS, Robertson AG, Schein JE, Siddiqui A, Smailus DE, Stott JM, Yang GS, Plummer F, Andonov A, Artsob H, Bastien N, Bernard K, Booth TF, Bowness D, Czub M, Drebot M, Fernando L, Flick R, Garbutt M, Gray M, Grolla A, Jones S, Feldmann H, Meyers A, Kabani A, Li Y, Normand S, Stroher U, Tipples GA, Tyler S, Vogrig R, Ward D, Watson B, Brunham RC, Krajden M, Petric M, Skowronski DM, Upton C, Roper RL.; ''The Genome sequence of the SARS-associated coronavirus.''; PubMedEurope PMCScholia
Huang C, Ito N, Tseng CT, Makino S.; ''Severe acute respiratory syndrome coronavirus 7a accessory protein is a viral structural protein.''; PubMedEurope PMCScholia
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
The PIK3C3-containing Beclin-1 complex consists of PIK3C3 (Vps34), BECN1 (Beclin-1, Atg6), PIK3R4 (p150, Vps15) and ATG14 (Barkor) (Matsunaga et al. 2009, Zhong et al. 2009). A similar complex where ATG14 is replaced by UVRAG functions later in autophagosome maturation and endocytic traffic (Itakura et al. 2008, Liang et al. 2008). Binding of KIAA0226 to this complex negatively regulates the maturation process (Matsunaga et al. 2009).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 DefinedSet instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
This COVID-19 Complex instance was generated via electronic inference from a curated CoV-1 (Human SARS coronavirus) Reactome instance. In Reactome, inference is the process used to automatically create orthologous Pathways, Reactions and PhysicalEntities from our expertly curated data (https://reactome.org/documentation/inferred-events).
Entry of influenza, parainfluenza and coronaviruses into airway epithelial cells requires binding of a viral spike protein to a host cell receptor, followed by cleavage and activation of the viral spike protein mediated by the host cell. Without this cleavage, fusion of the viral and host cell membranes is blocked. The primary receptor for the human SARS-CoV-1 virus is angiotensin converting enzyme 2 (ACE2) (Li et al. 2003). The resultant complex is cleaved by the protease transmembrane protease serine 2 (TMPRSS2) (Shulla et al. 2011, Heurich et al. 2014). Therefore, active site inhibitors of these airway proteases could have broad therapeutic applicability against multiple respiratory viruses (Laporte & Naesens 2017). The approved drug camostat is a protease inhibitor that may block SARS-CoV-2 entry into cells by inhibiting the actions of TMPRSS2 (Kawase et al. 2012, Hoffmann et al. 2020). Nafamostat, another serine protease inhibitor, was found to be a potent inhibitor of S-mediated membrane fusion and blocked MERS-CoV infection in vitro (Yamamoto et al. 2016).
Otamixaban (FXV673), an anticoagulant, is a potent and selective direct inhibitor of coagulation factor Xa. Virtual docking studies suggest that otamixaban may bind to the serine protease TMPRSS2 (Rensi et al. 2020, preprint). Inhibition of TMPRSS2 is being examined for antiviral activity but its inhibitory potential and/or antiviral activity have not yet been determined so it is annotated here as a candidate drug. I-432 is another inhibitor of TMPRSS2 under investigation for anti viral potential (Pászti-Gere et al. 2016).
Chloroquine (CQ) and hydroxychloroquine (HCQ) are diprotic weak bases that can exist in both protonated and unprotonated forms. Unprotonated CQ or HCQ can diffuse freely and rapidly across the membranes of cells and organelles to acidic cytoplasmic vesicles (late endosomes and lysosomes). Agents that have this ability are known as lysosomotropic agents. Once protonated, CQ2+ or HCQ2+ are trapped in the acidic lumen of these vesicles. This leads to an irreversible accumulation of CQ or HCQ in acidic vesicles to concentrations as much as 100 fold over cytosolic ones and to an elevation of vesicle pH due to trapping of H+ ions by CQ or HCQ. Thus, CQ analogues interfere with endosomal and lysosomal acidification, which in turn inhibits proteolysis, chemotaxis, phagocytosis and antigen presentation. As a result, cells are not able to proceed with endocytosis, exosome release and phagolysosomal fusion in an orderly manner (Foley & Tilley 1998, Yang & Shen 2020). In vitro, these endosomal acidification fusion inhibitors block cellular infection by a clinical isolate of SARS-CoV-2 (Wang et al. 2020, Hu et al. 2020).
Unprotonated chloroquine (CQ) and hydroxychloroquine (HCQ) can both diffuse freely and rapidly across the membranes of cells and organelles (Chinappi et al. 2010).
Lysosomes play critical roles in human biology receiving, trafficking, processing, and degrading biological molecules from cellular processes such as endocytosis, phagocytosis, autophagy and secretion. Lysosomes house around sixty proteolytic enzymes, among them cathepsins. Cathepsins are involved in many processes involving cell death, protein degradation, post-translational modifications of proteins, extracellular matrix (ECM) remodeling, autophagy, and immune signaling. The early stages of the viral life cycle involve the cleavage of the viral spike protein by cathepsin L (CTSL) in late endosomes, facilitating viral RNA release to continue viral replication. Teicoplanin, a glycopeptide antibiotic used to treat Gram-positive bacterial infection, especially in Staphylococcal infections, was shown to have efficacy in vitro against Ebola Virus, MERS and SARS-CoV-1 (Zhou et al. 2016).
Teicoplanin is thought to inhibit the low pH cleavage of the viral spike protein by CTSL in late endosomes thereby preventing the release of genomic viral RNA and the continuation of virus replication cycle (Baron et al. 2020). The target sequence that serve as the cleavage site for CTSL is conserved in the SARS-CoV-2 spike protein (Zhou et al. 2020 [preprint]). Further investigation is required to determine the therapeutic potential of teicoplanin in COVID-19 patients.
Relacatib is an investigational drug trialed for the treatment of osteoporosis (Duong et al. 2016). It is a potent CTSK inhibitor but also shows activity against CTSL (Kumar et al. 2007) so could potentially be investigated for Covid-19 patients. The antileprotic drug clofazimine and the antituberculous drugs rifampicin and isoniazid have been shown to inhibit cathepsins B, H and L from purified goat and bovine brains (Kamboj et al. 2003).
Inhibition of host cellular functions required for viral replication is considered another host-targeting antiviral strategy. Extensive pharmacological studies have validated ER glucosidases as valuable host antiviral targets against many enveloped viruses (Chang et al. 2013). Most known ER glucosidase inhibitors are imino sugars like 1-deoxynojirimycin (DNJ) and castanopermine (CAST) derivatives (Taylor et al. 1994).
It is generally believed that inhibition of ER glucosidase I and/or II prevents the removal of the terminal glucose moieties on N-linked glycans and results in misfolding and retention of glycoproteins in the ER and ultimate degradation via the ER-associated degradation (ERAD) pathway (Simsek et al. 2005, Alonzi et al. 2013). As a consequence of the abnormal trafficking and degradation of viral glycoproteins, virion assembly and secretion are inhibited (Chang et al. 2009, Taylor et al. 1998).
Long-term suppression of ER glucosidases I and/or II with more potent inhibitors may cause significant side effects, particularly in nerve and immune systems (Sadat et al. 2014).
Many GSK-3β inhibitors (GSKi) have been identified. They are known to induce apoptosis in leukemia and pancreatic cancer cells, and can destabilize p53, which may promote cellular death in response to DNA damaging agents (Wang et al, 2008; Beurel et al, 2009). Administration of GSKi inhibited cochlear destruction in cisplatin-injected mice (Park et al, 2009).
Li is a selective ATP competitive inhibitor of GSK-3 (Ryves and Harwood, 2001). Lithium carbonate has been and continously is in clinical trials with bipolar disorder patients (Moore et al, 2009). LY2090314 has been in clinical trials for metastatic pancreatic cancer and acute leukemia ([NCT01632306], [NCT01287520], [NCT01214603]). Clinical trials of GSKi for Alzheimer's disease were unsuccessful.
The use of GSKi remains controversial because of their possibly oncogenic properties. Evaluation of GSKi in clinical trials has been hampered by the fear that inhibition of GSK-3 may stimulate or aid in malignant transformation as GSK-3 can phosphorylate pro-oncogenic factors such as beta-catenin, c-Jun and c-Myc which targets them for degradation (Patel & Woodgett, 2008). However, no studies have been reported suggesting that treatment of mice with GSKi resulted in an increase in cancer incidence. In fact, many patients with bi-polar disorder have been treated with lithium for prolonged periods of time. There does not appear to be any evidence that these patients have increased incidences of cancer (McCubrey et al, 2014).
The GSKi kenpaullone and lithium chloride were found to reduce viral Nucleoprotein phosphorylation in the severe acute respiratory syndrome CoV-infected VeroE6 cells and decrease the viral titer and cytopathic symptoms. Effect of GSK-3 inhibition were reproduced in another coronavirus, the neurotropic JHM strain of mouse hepatitis virus (Wu et al, 2009).
SARS-CoV-2 nonstructural proteins nsp7 and nsp8 form a heterodimer (Gao et al. 2020, Li et al. 2020). A dimer of dimers has also been observed and it has been proposed that this heterotetramer forms a putative RNA-binding site (Konkolova et al. 2020).
The nsp7:nsp8 heterodimer binds to the RNA-directed RNA polymerase (nsp12) of the SARS-CoV-2 (Gao et al. 2020, Li et al. 2020, Hillen et al. 2020). The second subunit of nsp8, not bound to nsp7, interacts with a different region of nsp12 (Gao et al. 2020, Yin et al. 2020), as previously found in SARS-CoV-1 (Kirchdoerfer and Ward 2019).
Nonstructural protein 9 from SARS-Cov-2 is an obligate homodimer, comprising a unique fold that associates via an unusual α-helical GxxxG interaction motif. The integrity of this motif is considered important for viral replication (Littler et al, 2020; Miknis et al, 2009).
nsp13 of SARS-CoV-2 possesses the nucleoside triphosphate hydrolase (NTPase) activity (Chen et al. 2020, Shu et al. 2020), and functions as an NTP-dependent RNA helicase that can unwind RNA helices (Shu et al. 2020). Several G-quadruplex structures were confirmed in SARS-CoV-2 RNA and found to directly interact with nsp13, which may act to melt these structures (Ji et al. 2020).
nsp13 of SARS-CoV-1 is an ATP-dependent helicase that functions in the 5'-3' direction to unwind double stranded RNAs that have a 5' single strand overhang at least 20 nucleotides long. nsp13 can also act on double strand DNA in vitro, but dsRNA is thought to be its physiological substrate. The catalytic activity of SARS-CoV-1 nsp13 is increased in the presence of nsp12, the viral RNA-dependent RNA polymerase. nsp13 is needed for the replication of SARS-CoV-1 and is thought to act by melting secondary structures in the genomic RNA template during replication, and also to be involved in unwinding of RNA duplexes during transcription of viral genes. nsp13 is a promising target for experimental anti-SARS-CoV-1 drugs (Tanner et al. 2003, Ivanov et al. 2004, Bernini et al. 2006, Chen et al. 2009, Lee et al. 2010, Adedeji et al. 2012).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
In about 15% of translation attempts of the genomic viral mRNA1 in the host cell cytosol, a -1 frameshift happens after the nsp10 gene that leads to translation of the full pp1ab polyprotein (Baranov et al, 2005).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
nsp8 functions as an RNA-dependent RNA polymerase (RdRp) that serves as the primase for nsp12, the main RdRp of the SARS coronavirus 1 (SARS-CoV-1) (Imbert et al. 2006), as it is capable of de novo RNA synthesis (te Velthuis et al. 2011). nsp8 synthesizes short oligonucleotides (less than 6 bases long) using genomic RNA as a template. nsp8 requires at least one cytidine residue in the template sequence for its activity. Activity is dependent on manganese ions (Imbert et al. 2006). nsp8 can also extend primers but is 20-fold less efficient than nsp12 (te Velthuis et al. 2011).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Based on studies in other coronaviruses, the final SARS-COV-2 ribonucleoprotein complex is predicted to be a hollow helical structure with an approximate diameter of 9-16nm, with the C-terminal domain of N protein forming the inner core and the N-terminal domain forming the outer surface (Neuman et al, 2006; Chen et al, 2007). Oliomerization of the N protein capsid coat is likely nucleated through both protein-RNA and protein-protein interactions by the first few N-protein dimers on the genomic RNA (Saikatendu et al, 2007; Chang et al, 2013; reviewed in Chang et al, 2014). Each N dimer may make contact with up to 7 bases of the RNA (reviewed in Chang et al, 2014).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Within the host cell endocytic vesicle, SARS-CoV-2 Spike (S) protein is cleaved between residues 797 and 798 by cathepsin L1 (CTSL) (Huang et al. 2006). The roles of S protein in viral binding to the host cell membrane and fusion of viral and host cell membranes and thus the central role of S protein in determining the host range and tissue tropisms of the virus are reviewed by Belouzard et al. (2012).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
The majority of nucleoprotein is serine-phosphorylated in the cytosol and, possibly, in the nucleus where it gets immediately transported to the cytosol. Phosphorylation is catalyzed by glycogen synthase kinase 3 (GSK3) and several other host cell kinases (Surjit et al, 2005; Wu et al. 2009)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Both a predicted beta-hairpin motif and the N-terminal part of protein E are sufficient for its localization to the Golgi membrane. Although porin activity has been shown for protein E it cannot be detected in the plasma membrane of infected cells (Liao et al, 2006; Cohen et al, 2011; Nieto-Torres, 2011).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Non-structural protein 14 (nsp14) of the human SARS coronavirus is a bifunctional enzyme bearing 3'-5 exoribonuclease activity involved in replication fidelity and RNA cap N7-guanine methyltransferase activity involved in 5'-RNA capping. nsp14 binds to the minimal replication and transcription complex (RTC), composed of nsp7, nsp8, and nsp12, by directly binding to nsp12 (the main RNA-dependent RNA polymerase). Binding of nsp14 does not affect the processivity of the RTC (Minskaia et al. 2006, Subissi et al. 2014).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Membranous structures containing protein 3a are being shedded from the cell membrane (Huang et al, 2006)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
nsp3 and nsp4 alone caused considerable ER membrane deformation, producing a perinuclear double-walled maze-like body (MLB), and the nsp3–nsp4 interaction was shown to be absolutely necessary for such membrane rearrangement. Further necessary factors are nsp6 and unidentified host factors (Angelini et al 2013, Sakai et al 2017). nsp6 by itself can form Atg5 and LC3II-positive vesicles classically observed in autophagy (Cottam et al, 2014). However, in mouse hepatitis virus (MHV) infections, EDEM1 and OS9 of the ER-associated degradation system have been shown to be necessary co-factors (Reggiori et al, 2010)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
To process the nsp3/4 cleavage site, PL-PRO and, presumably, nsp3-4 need to be glycosylated and localized to a membrane (Harcourt et al, 2004)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
In most translation attempts the genomic viral mRNA1 in the cytosol is translated to a shortened polyprotein, pp1a, that does not contain genome replication enzymes (Baranov et al, 2005).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Calnexin transiently binds the unfolded spike protein and prevents its aggregation and premature degradation, ensuring its correct folding (Fukushi et al, 2012).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
The papain-like protease domain of the nsp1-4 fragment alone is sufficient for processing the nsp1/2 and nsp2/3 cleavage sites (Harcourt et al, 2004).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Two of the four cysteine-rich clusters of the viral spike protein are modified by palmitoylation. This is required for the protein's partitioning into detergent-resistant membranes and for cell–cell fusion. In general, palmitoylation is usually non-enzymatic (Petit et al, 2007; McBride and Machamer, 2010; Veit, 2012).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
SARS-CoV-1 encodes eight subgenomic RNAs, mRNA2 to mRNA9. mRNA1 corresponds to the genomic RNA. The 5' and 3' ends of subgenomic RNAs are identical, in accordance with the template switch model of coronavirus RNA transcription (Snijder et al. 2003, Thiel et al. 2003, Yount et al. 2003). Therefore, consistent with this and the studies of the murine hepatitis virus (MHV), which is closely related to SARS-CoV-1, genomic positive strand RNA is first transcribed into negative sense (minus strand) subgenomic mRNAs, that subsequently serve as templates for the synthesis of positive strand subgenomic mRNAs. Negative-sense virus RNAs are present in much smaller amounts than positive-sense RNAs (Irigoyen et al. 2016). Each subgenomic RNA contains a leader transcription regulatory sequence (leader TRS) that is identical to the leader of the genome, appended via polymerase “jumping� during negative strand synthesis to the body transcription regulatory sequence (body TRS), a short, AU-rich motif of about 10 nucleotides found upstream of each ORF that is destined to become 5' proximal in one of the subgenome-length mRNAs. The 3' and 5'UTRs may interact through RNA–RNA and/or RNA–protein plus protein–protein interactions to promote circularization of the coronavirus genome, placing the elongating minus strand in a favorable topology for leader-body joining. The host protein PABP was found to bind to the coronavirus 3' poly(A) tail and to interact with the host protein eIF-4G, a component of the three-subunit complex that binds to mRNA cap structures, which may promote the circularization of the coronavirus genome. Two viral proteins that bind to the coronavirus 5'UTR, the N protein and nsp1, may play a role in template switching. The poly(A) tail is necessary for the initiation of minus-strand RNA synthesis at the 3' end of genomic RNA. For review, please refer to Sawicki et al. 2007 and Yang and Leibowitz 2015.
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
A certain part of the nucleoprotein can be found in the nucleolus. This localisation seems to depend on the protein's sumoylation (Li et al, 2005)
N-glycan side chains on the unfolded SARS-CoV-2 spike protein get their terminal glucose moieties cleaved by ER glucosidases I and II, before folding. (Watanabe et al, 2020, Völker et al, 2002, Pelletier et al, 2000).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Protein M accumulates in the Golgi complex and recruits Spike protein to the sites of virus assembly and budding in the ERGIC (Voss et al, 2009).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Significant amounts of the unphosphorylated N protein are associated with the cell membrane (Surjit et al, 2005)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
The crucial step of autocleavage of pp1a involves the formation of an "intermediate" pp1a dimer which has weak protease activity. This "embedded" 3CLp liberates itself by cleaving the ends off its monomer in trans. Only after that occurs does the cleaved 3CLp form a dimer, the most efficient form of the enzyme (Hsu et al, 2005; Chen et al, 2010; Muramatsu et al, 2016)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
To process the nsp3/4 cleavage site, PL-PRO and, presumably, nsp3-4 need to be glycosylated and localized to a membrane (Harcourt et al, 2004)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Human SARS coronavirus 1 (SARS-CoV-1) non-structural protein 15 (nsp15) contains the LXCXE/D motif characteristic of proteins that bind to the retinoblastoma protein RB1. Binding to human RB1 increases the endonuclease activity of nsp15 but is not required for it. RB1 bound to nsp15 is retained in the cytosol. Interaction of nsp15 with RB1 likely affects the cell cycle of infected cells and probably modulates cytotoxicity of SARS-CoV-1 (Bhardwaj et al. 2012).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Viral protein 3a translocates from the cytosol to the ERGIC (endoplasmic reticulum Golgi intermediate compartment) (Oostra et al. 2006)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Viral E protein is modified by palmitoylation at all three cysteine residues. In general, palmitoylation is usually non-enzymatic (Liao et al, 2006, Veit, 2012).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
nsp13, the helicase of the human SARS coronavirus 1 (SARS-CoV-1) binds to DDX5, a host protein implicated in transcription, pre-mRNA processing, RNA degradation, RNA export, ribosome assembly and translation. DDX5 knockdown inhibits viral replication (Chen et al. 2009).
Remdesivir (GS-5734) is an investigational nucleotide analogue drug that was developed for its broad spectrum antiviral potential against Ebola and Marburg virus activity (Siegel et al. 2017). It targets and inhibitis viral RNA-dependent RNA polymerase (nsp12, RdRP), the key component of the replication transcription complex (RTC) (Agostini et al. 2018, Brown et al. 2019, Gordon et al. 2020). Remdesivir is being investigated for potential antiviral activity against SARS-CoV-2 by targeting viral replication (Agostini et al. 2018). Gordon et al. demonstrate remdesivir possesses broad antiviral activity against RNA viruses, including SARS-CoV, MERS-CoV and SARS-CoV-2 in-vitro (Gordon et al. 2020b). It could prevent asymptomatic, mild or moderate Covid-19 cases from progressing to severe disease (clinical trials NCT04252664, NCT04257656) but results so far in infected people have been mixed.
EIDD-2801, is an isopropylester prodrug of the ribonucleoside analogue N4-hydroxycytidine (NHC, EIDD-1931) that shows broad spectrum antiviral activity against various RNA viruses including Ebola, Influenza and CoV (Toots et al. 2019). NHC acts as a competitive alternative substrate for virally encoded RNA-dependent RNA polymerases. NHC was shown to inhibit multiple genetically-distinct Bat-CoV viruses in human primary epithelial cells without affecting cell viability. Prophylactic/therapeutic oral administration of NHC reduced lung titers and prevented acute lung failure in C57B/6 mice infected with CoV. The potency of NHC against multiple coronaviruses, its therapeutic efficacy, and oral bioavailability in vivo, all highlight its potential as an effective antiviral against SARS-CoV-2 in Covid-19 infections and other future zoonotic coronaviruses (Sheahan et al. 2020).
nsp15 of SARS-CoV-2 shares 88% sequence identity and 95% sequence similarity with nsp15 of SARS-CoV-1. Similar to its SARS-CoV-1 orthologue, nsp15 of SARS-CoV-2 forms a hexamerthat consists of a dimer of trimers. Hexamer formation is necessary for the endonuclease activity of nsp15. nsp15 preferentially cleaves 3' of uridines, generating 2'-3' cyclic phosphates after cleavage. nsp15 requires Mn2+ ions for catalytic activity. C-terminal domain contains the active site, which faces away from the center of the hexamer and contains the extreme C-terminal residues (Kim et al. 2020).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
A sialyltransferase adds a terminal sialic acid moiety to protein 3a with an O-linked glycosyl side chain. This glycosylated form later is associated with the virion (Oostra et al, 2006)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Main protease cleaves all cleavage sites of pp1a and ppa1b starting with nsp4/5, thus cleaving itself, and all the cytosolic RTC proteins (Fan et al, 2004)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
The ribonucleoprotein complex is recruited to the assembling virion through interactions with the M C-terminal tail, and appears to be independent of viral RNA (Hsieh et al, 2008; Hatakeyama et al, 2008; He et al, 2004; Luo et al, 2006; reviewed in Ujike and Taguchi, 2015). How much of this is fully conserved in SARS-COV-2 remains to be experimentally verified.
The interaction between the non-structural proteins nsp16 and nsp10 is conserved in SARS-CoV-2 virus (Li et al. 2020, Viswanathan et al. 2020, Rosas-Lemus et al. 2020).
In SARS-CoV-1, nsp16 was identified as an AdoMet-dependent (nucleoside-2'O)-methyltransferase involved in capping of viral RNAs. nsp16 binds to nsp10, which serves as a cofactor for nsp16 (Bouvet et al. 2010, Lugari et al. 2010). Nsp16 alone is unstable and exhibits 2'-O-methyltransferase activity only in complex with nsp10 (Debarnot et al, 2011; Decroly et al, 2011). nsp10-mediated activation of nsp16 catalytic activity is conserved in all coronaviruses (Wang et al. 2015). The same binding surface of nsp10 interacts with nsp14 and nsp16, suggesting that binding of nsp14 and nsp16 to nsp10 is mutually exclusive. However, as nsp10 is produced in a higher number of copies than nsp14 and nsp16, and as nsp14 and nsp16 act coordinately in RNA capping, it is most likely that nsp14:nsp10 and nsp16:nsp10 complexes co-exist within the viral replication-transcription complex (RTC) (Bouvet et al. 2012, Bouvet et al. 2014). One structural study reported that nsp10 forms dodecamers (Su et al. 2006), which would potentially allow simultaneous binding of nsp14 and nsp16 to nsp10 homomeric complexes, but it is not certain if such homomeric complexes of nsp10 exist in vivo, and if the structure of the nsp10 dodecamer would be permissive for nsp16 binding (Chen et al. 2011). nsp10 contains two zinc fingers which are thought to be involved in RNA binding (Su et al. 2006, Joseph et al. 2006).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Protein E forms a pentamer of monomers without disulfide bonds (Parthasarathy et al, 2012)
The cryo-electron microscopy (cryoEM) structure of the SARS-CoV-2 replication transcription complex (RTC) components nsp7, nsp8 and nsp12, bound to more than two turns of RNA template-product duplex, indicates that the active cleft of nsp12 binds to the first turn of RNA, while two copies of nsp8 bind to opposite sides of the cleft and position the second turn of RNA. Long helical extensions in nsp8 protrude along exiting RNA, forming positively charged "sliding poles". These sliding poles may confer processivity to the RTC (Hillen et al. 2020). Binding of nsp12 to the template RNA is markedly increased by the presence of nsp7 and nsp8 (Yin et al. 2020). Notable structural rearrangements occur in nsp12, nsp8 and nsp7 to accommodate the RNA (Wang et al. 2020).
Based on studies in SARS-CoV-1, the RTC binds to the 3' end of the viral plus strand genomic RNA to initiate synthesis of the complementary minus strand. A 36 nucleotide sequence from the 3’-UTR of the plus strand, predicted to form a stable stem-loop structure, seems to be the minimal cis-acting RNA element required for the viral RNA-directed RNA polymerase (nsp12) to initiate RNA synthesis. The polyA tail also seems to play a role in the initiation of replication of viral genomic RNA (Ahn et al. 2012).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Binding of the genomic RNA to one or a small number of N-N dimers may be the initiating event in nucleocapsid formation. Both the NTD and the CTD of N have been shown to have RNA-binding activity (Huang et al, 2004a; Huang et al, 2004b; Chen et al, 2007; Chang et al, 2009; Takeda et al, 2008), and the IDRs seem likely to also contribute (Chang et al, 2009). These initial binding events may nucleate nucleocapsid formation through further recruitment of N protein dimers (reviewed in Chang et al, 2014).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
N protein is sumoylated at a lysine residue. Abolition of sumoylation of nucleoprotein significantly decreases homo-oligomerisation of the protein (Li et al, 2005)
SARS-CoV-2 plus strand genomic RNA (gRNA), like genomic RNAs of SARS-CoV-1 and other coronaviruses, is polyadenylated. The poly(A) tail of gRNA is longer than the poly(A) tail of subgenomic SARS-CoV-2 RNAs (Kim et al. 2020). Coronavirus plus strand gRNAs possess a polyadenylation signal in their 3'UTR and are polyadenylated by an undetermined viral RNA polymerase, possibly nsp8 or nsp12 (Spagnolo and Hogue 2000, Peng et al. 2016, Tvarogova et al. 2019).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
The genomic and subgenomic (sg) mRNAs of SARS-CoV-1 coronavirus are presumed to be capped at their 5′ end, based on studies of the mouse hepatitis virus (MHV) (Lai and Stohlman 1981) and the equine torovirus (van Vliet et al. 2002). Non-structural protein 14 (nsp14) acts as an RNA guanine-N7-methyltransferase (N7-MTase) that completes the synthesis of the cap-0 on SARS-CoV-1 mRNAs. The cap-0 represents N7-methyl guanosine connected to the 5′ nucleotide through a 5′ to 5′ triphosphate linkage, and is also known as m7G cap or m7Gppp cap. The N7-MTase domain maps to the carboxy-terminal part of nsp14 (Chen et al. 2009). Cap-0 formation requires three sequential reactions catalyzed by RNA triphosphatase (TPase), guanylyltransferase (GTase), and N7-MTase. There is no evidence that nsp14 possesses TPase and GTase activities, and no other SARS-CoV-1 proteins with these activities have been identified, so the identities of the enzymes that mediate these required steps remain unknown. Based on the study of the human coronavirus 229E, non-structural protein 13 (nsp13) may have a TPase activity in addition to its established helicase activity (Ivanov and Ziebuhr 2004).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Despite its importance, expression of M alone is not sufficient to drive formation of a mature virus (reviewed in Masters, 2006). Protein-protein interactions between M and S, N and E, among other components, are required for assembly of a mature virus and for membrane curvature. Many studies have examined the minimal system required for release of viral like particles (VLPs) with sometimes contradictory results, but interactions between M, N and E are sufficient to promote release of significant numbers of VLPs (Ho et al, 2004; Huang et al, 2004; Mortola and Roy, 2004; Hsieh et al, 2005; Siu et al, 2008; Hatakeyama et al, 2008; Tseng et al, 2013; reviewed in Masters, 2006)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
The genomic and subgenomic mRNAs of SARS-CoV-1 coronavirus, including the minus strand genomic RNA complement, are presumed to be capped at their 5′ end, based on studies of the mouse hepatitis virus (MHV) (Lai and Stohlman 1981) and the equine torovirus (van Vliet et al. 2002). The non-structural protein 14 (nsp14) acts as an RNA guanine-N7-methyltransferase (N7-MTase) that completes the synthesis of the cap-0 on SARS-CoV-1 minus strand genomic RNA. The cap-0 represents N7-methyl guanosine connected to the 5′ nucleotide through a 5′ to 5′ triphosphate linkage, and is also known as m7G cap or m7Gppp cap. The N7-MTase domain maps to the carboxy-terminal part of nsp14 (Chen et al. 2009). Cap-0 formation requires three sequential reactions catalyzed by RNA triphosphatase (TPase), guanylyltransferase (GTase), and N7-MTase. There is no evidence that nsp14 possesses TPase and GTase activities, and no other SARS-CoV-1 proteins with these activities have been identified, so the identities of the enzymes that mediate these required steps remain unknown. Based on the study of the human coronavirus 229E, non-structural protein 13 (nsp13) may have a TPase activity in addition to its established helicase activity (Ivanov and Ziebuhr 2004).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
After synthesizing the complementary minus RNA of the plus strand viral genomic RNA, SARS-CoV-1 replication-transcription complex (RTC) associates with the minus strand to initiate plus strand synthesis and to initiate transcription of subgenomic (sg) mRNAs (Ahn et al. 2012).
The function of the non-structural protein nsp16 as a 2'O-methyltransferase that acts in complex with its co-activator nsp10 is conserved in SARS-CoV-2 (Viswanathan et al. 2020).
The subgenomic mRNAs of SARS-CoV-2 coronavirus are presumed to be capped at their 5′ ends, based on studies of the mouse hepatitis virus (MHV) (Lai and Stohlman 1981) and the equine torovirus (van Vliet et al. 2002). The non-structural protein 16 (nsp16) acts as a 2'O-methyltransferase that converts coronavirus cap-0 to cap-1, which was first demonstrated with nsp16 cloned from the feline coronavirus (FCV) (Decroly et al. 2008). Cap-0 represents N7-methyl guanosine connected to the 5′ nucleotide through a 5′ to 5′ triphosphate linkage (also known as m7G cap or m7Gppp cap). Cap-1 is generated by an additional methylation on the 2′O position of the initiating nucleotide, and is also known as m7GpppNm. The non-structural protein 10 (nsp10) acts as an activator of nsp16 and is necessary for cap-1 synthesis (Bouvet et al. 2010, Decroly et al. 2011). Coronavirus RNAs with cap-1 are protected from IFIT-mediated interferon response, as IFITs recognize unmethylated 2'-O RNAs. IFITs are interferon-induced proteins with tetratricopeptide repeats that recognize unmethylated 2'-O RNAs and act to inhibit expression of virally encoded mRNAs (Menachery et al. 2014).
SARS-CoV-2 produces nine subgenomic RNAs (sgRNAs). N protein-encoding mRNA (mRNA9) is most abundantly expressed, followed by mRNAs encoding proteins S (mRNA2), 7a (mRNA7a), 3a (mRNA3), 8 (mRNA8), M (mRNA5), E (mRNA4), 6 (mRNA6) and 7b (mRNA7b) (Kim et al. 2020).
SARS-CoV-1 encodes eight subgenomic RNAs, mRNA2 to mRNA9. mRNA1 corresponds to the genomic RNA. mRNA2 encodes the S protein. mRNA3 is bicistronic and encodes proteins 3a and 3b. mRNA4 encodes the E protein. mRNA5 encodes the M protein. mRNA6 encodes the protein 6. mRNA7, mRNA8 and mRNA9 are bicistronic, with mRNA7 encoding proteins 7a and 7b, mRNA8 encoding proteins 8a and 8b, and mRNA 9 encoding proteins 9a and N. The 5' and 3' ends of subgenomic RNAs are identical, in accordance with the template switch model of coronavirus RNA transcription (Snijder et al. 2003, Thiel et al. 2003, Yount et al. 2003). Based on studies of the murine hepatitis virus (MHV), which is closely related to SARS-CoV-1, positive-sense virus mRNAs are present at much higher amounts than negative-sense mRNAs (Irigoyen et al. 2016).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Human von Hippel Lindau (VHL) protein, a tumor suppressor that acts as a component of an E3 ubiquitin ligase complex, interacts with the non-structural protein 16 (nsp16) of the human SARS coronavirus 1 (SARS-CoV-1) and the mouse hepatitis virus, also a coronavirus. VHL negatively regulates SARS-CoV-1 replication, but the exact mechanism is not known (Yu et al. 2015).
The function of the non-structural protein nsp16 as a 2'O-methyltransferase that acts in complex with its co-activator nsp10 is conserved in SARS-CoV-2 (Viswanathan et al. 2020).
The genomic and subgenomic mRNAs of SARS-CoV-2 coronavirus, including the minus strand genomic RNA, are presumed to be capped at their 5′ end, based on studies of the mouse hepatitis virus (MHV) (Lai and Stohlman 1981) and the equine torovirus (van Vliet et al. 2002). Non-structural protein 16 (nsp16) acts as a 2'O-methyltransferase that converts coronavirus cap-0 to cap-1, which was first demonstrated with nsp16 cloned from the feline coronavirus (FCV) (Decroly et al. 2008). Cap-0 represents N7-methyl guanosine connected to the 5′ nucleotide through a 5′ to 5′ triphosphate linkage (also known as m7G cap or m7Gppp cap). Cap-1 is generated by an additional methylation on the 2′O position of the initiating nucleotide, and is also known as m7GpppNm. Non-structural protein 10 (nsp10) acts as an activator of nsp16 and is necessary for cap-1 synthesis (Bouvet et al. 2010, Decroly et al. 2011). Coronavirus RNAs with cap-1 are protected from IFIT-mediated interferon response. IFITs are interferon-induced proteins with tetratricopeptide repeats that recognize unmethylated 2'-O RNAs and act to inhibit expression of virally encoded mRNAs (Menachery et al. 2014).
nsp13, which functions as the viral helicase, is as a part of the SARS coronavirus 2 (SARS-CoV-2) replication-transcription complex (RTC). Two molecules of SARS-CoV-2 nsp13 are found in the RTC complex. N-terminal domains of each nsp13 interact with N-terminal extensions of each copy of nsp8, while nsp13 molecule interacts with the thumb region of nsp12, the viral RNA-dependent RNA polymerase. One nsp13 molecule of SARS-CoV-2 is tightly bound to the RTC, while the other nsp13 molecule is dissociable (Chen et al. 2020). In SARS-CoV-1, nsp13 has been reported to directly interact with nsp12, and nsp12 was shown to increase the helicase activity of nsp13 (von Brunn et al. 2007, Adedeji et al. 2012, Jia et al. 2019).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Protein M is exclusively N-glycosylated at one asparagine by an unknown glycosyltransferase. However, further processing of N-linked glycans is prevented in infected cells. Both the glycosylated and nonglycosylated M is incorporated into the virion. In summary, glycosylation of M is neither a prerequisite for intra-cellular transport nor for recruitment into the virion (Voss et al, 2006).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
In addition to the main structural proteins and the nucleocapsid, the mature virion may also contain low proportions of accessory proteins, including protein 3a and 7a (reviewed in McBride and Fielding, 2012). Protein 3a has been shown to interact with E, M, S and protein 7a and is estimated to be present in the virion at 2/3 the molar ratio of E protein (Ito et al, 2005; Shen et al, 2005; Tan et al, 2004). Although 3a tetramers are predicted to act as ion channels in the host plasma membrane, increasing cell permeability, the role of 3a in the mature virion is not clear (Lu et al, 2006; reviewed in McBride and Fielding, 2012). Protein 7a is type 1 transmembrane protein that interacts with M, E, S and protein 3a and may be incorporated into the mature virion. What functional role protein 7a may play in the assembled virion is unclear (Huang et al, 2006; Fielding et al, 2004; Tan et al, 2004).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Viral E protein is ubiquitinated both in vitro and in cells (Alvarez et al, 2011).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Nonstructural protein 15 (nsp15) of the SARS coronavirus (SARS-CoV-1) binds to the replication-transcription complex (RTC) through interaction with nsp8 (Imbert et al. 2008). This interaction appears to be conserved in other coronaviruses, such as mouse hepatitis virus (MHV) (Athmer et al. 2017). nsp15 is an endonuclease characteristic for the order Nidovirales that includes the family Coronaviridae. nsp15 preferentially cleaves 3' of uridines, generating 2'-3' cyclic phosphates after cleavage. nsp15 requires Mn2+ ions for catalytic activity. Functional nsp15 is needed for production of viable virions and for viral transcription (Guarino et al. 2005, Ricagno et al. 2006, Bhardwaj et al. 2006, Joseph et al. 2007, Bhardwaj et al. 2008, Bhardwaj et al. 2012). The biological role of nsp15 has not been elucidated. It may degrade host mRNAs to shut down host translation, but so far no human or viral RNA targets have been identified.
Remdesivir (GS-5734) is an investigational nucleotide analogue drug that was developed for its broad spectrum antiviral potential against Ebola and Marburg virus activity (Siegel et al. 2017). It targets and inhibitis viral RNA-dependent RNA polymerase (nsp12, RdRP), the key component of the replication transcription complex (RTC) (Agostini et al. 2018, Brown et al. 2019, Gordon et al. 2020). Remdesivir is being investigated for potential antiviral activity against SARS-CoV-2 by targeting viral replication (Agostini et al. 2018). Gordon et al. demonstrate remdesivir possesses broad antiviral activity against RNA viruses, including SARS-CoV, MERS-CoV and SARS-CoV-2 in-vitro (Gordon et al. 2020b). It could prevent asymptomatic, mild or moderate COVID-19 cases from progressing to severe disease (clinical trials NCT04252664, NCT04257656) but results so far in infected people have been mixed.
EIDD-2801, is an isopropylester prodrug of the ribonucleoside analogue N4-hydroxycytidine (NHC, EIDD-1931) that shows broad spectrum antiviral activity against various RNA viruses including Ebola, Influenza and CoV (Toots et al. 2019). NHC acts as a competitive alternative substrate for virally encoded RNA-dependent RNA polymerases. NHC was shown to inhibit multiple genetically-distinct Bat-CoV viruses in human primary epithelial cells without affecting cell viability. Prophylactic/therapeutic oral administration of NHC reduced lung titers and prevented acute lung failure in C57B/6 mice infected with CoV. The potency of NHC against multiple coronaviruses, its therapeutic efficacy, and oral bioavailability in vivo, all highlight its potential as an effective antiviral against SARS-CoV-2 and other future zoonotic coronaviruses (Sheahan et al. 2020).
The interaction between nsp10 and nsp14 is conserved in SARS-CoV-2 virus (Li et al. 2020). In SARS-CoV-1, nsp10 was shown to form a stable complex with nsp14 (Bouvet et al. 2012) and serve as a co-factor for nsp14, stimulating its 3'->5' exonuclease activity (Bouvet et al. 2012, Subissi et al. 2014, Bouvet et al. 2014).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
The replication-transcription complex (RTC) completes synthesis of the genomic RNA complement (minus strand). The complex of nsp7 and nsp8 confers processivity to nsp12, the virally encoded RNA-dependent RNA polymerase that replicates the viral genomic RNA, enabling the RTC to complete the RNA synthesis with a very low dissociation rate. nsp7 plays a crucial role in maintaining binding of the RTC to the RNA. nsp14 subunit of the RTC does not affect the processivity (Subissi et al. 2014).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Main protease cleaves all cleavage sites of pp1a and ppa1b starting with nsp4/5, thus cleaving itself, and all the cytosolic RTC proteins (Fan et al, 2004)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Based on studies in other coronaviruses, SARS-COV-2 S trimers are presumed to be recruited to the assembling virion through interaction with M protein (reviewed in Ujike and Taguchi, 2015). Multiple regions of M contribute to the recruitment of S, with a single tyrosine residue in the C-terminal domain of M playing a critical role (McBride et al, 2010a; Hsieh et al, 2008). Interaction with M is aided by a dibasic motif in the C-terminus of S, which promotes retrieval of the spike protein from the cell surface by binding the COPI coat (McBride et al, 2007; Ujike et al, 2016). Palmitoylation of the C-terminus of S appears dispensible for the interaction with M in SARS-COV-1, unlike the case in other coronaviruses; whether this is also true for SARS-COV-2 remains to be determined (Ujike et al, 2012; McBride 2010b; reviewed in Ujike and Taguchi, 2015). Size estimates and modelling suggest the mature virion has approximately 300 S trimers (Neuman et al, 2006; reviewed in Chang et al, 2014).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Protein M accumulates in the Golgi complex and recruits Spike protein to the sites of virus assembly and budding in the ERGIC (Voss et al, 2009).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
In the host cell cytosol the pp1a polyprotein spontaneously dimerizes. This temporary dimer has weak protease activity (Chen et al, 2010)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
As early as 3 hours post-infection, cytoplasmic accumulations of N are formed in infected cells, they colocalize with viral RNA. From 5 hours post-infection on, N can be detected in the Golgi, the budding site (Stertz et al, 2007)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
As it contains cargo sorting motifs in its cytoplasmic domain, protein 3a gets localized by the cell's protein transport system to the cell membrane where it functions as an ion channel (Tan et al, 2004). This ion channel function is necessary for the protein's pro-apoptotic function (Chan et al, 2009)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Some phosphorylated N is found to associate with the cell membrane (Surjit et al, 2005).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
On the final assembly, proteomics data suggest that the NAB−βSM−TM1 domains of nsp3 can interact with nsp7 − 8, as well as nsps 12–16, and the domain Y1 plus CoV-Y interacts with nsp9 and nsp12 (Imbert et al., 2008). Also a PL2pro−NAB−βSM−TM1 construct of Nsp3 can bind Nsp4 and Nsp12, while the region from TM1 to the end of Nsp3 only binds Nsp8 (Pan et al., 2008).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
SARS-CoV-2 spike (S3a) protein, as a component of the S3:M:E:encapsidated SARS coronavirus genomic RNA: 7a:O-glycosyl 3a tetramer complex, binds to glycosylated angiotensin converting enzyme 2 (ACE2) associated with the human host cell plasma membrane. Structural studies of the interaction between human ACE2 protein and the receptor-binding domain of S3a protein have identified key amino acid residues in both proteins responsible for their high-affinity interaction. These residues may be a key factor determining severity (and possibly human-to-human transmission) of SARS-CoV-2 (Li et al. 2003, 2005). The roles of S protein in viral binding to the host cell membrane and fusion of viral and host cell membranes and thus the central role of S protein in determining the host range and tissue tropisms of the virus are reviewed by Belouzard et al. (2012).
This COVID‑19 event has been created by a combination of computational inference from SARS-CoV-1 data (https://reactome.org/documentation/inferred-events) and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
The replicase polyprotein 1a of the human severe acute respiratory syndrome coronavirus is post-translationally cleaved by virally encoded proteases to generate non-structural proteins (nsps). Viral nsps induce the formation of ER-bound double membrane vesicles (DMV) in host cells post infection. These DMVs are decorated with host microtubule-associated proteins 1A/1B light chain 3B (MAP1LC3B) proteins that are involved in autophagosome formation. However, there is no evidence that DMVs are recruited to the autophagy machinery. Immunofluorescence studies show that nsp8 colocalizes with MAP1LC3B suggesting a binding event (Prentice E. et al 2004).
After synthesizing the complementary minus RNA of the plus strand viral genomic RNA, virally encoded RNA-dependent RNA polymerase (nsp12, also known as RdRP) uses the minus strand as a template to generate viral genomic RNA that can be packaged into virions. SARS-CoV-2-derived nsp12, in complex with nsp7 and nsp8, was shown to have RNA polymerization activity on a poly-U template (Yin et al. 2020). Details of SARS-CoV-2 replication have not yet been elucidated and are inferred from SARS-CoV-1. Purified SARS-CoV-1 nsp12 shows both primer dependent and primer-independent RNA synthesis activity in vitro. nsp12 is able to initiate RNA synthesis with as little as 37 nucleotides of RNA from the 3’ end of the minus strand viral RNA (complementary to the 5’-UTR of the plus strand genomic RNA - c5’-UTR). Similar to the 3'-UTR of the plus strand, the 3' end of the minus strand (c5’-UTR) is predicted to form a stable stem-loop structure and seems to be the minimal cis-acting RNA element required for nsp12 to initiate RNA synthesis using the minus strand as a template (Ahn et al. 2012). It is unclear if replication of the minus strand is primer-dependent. The complex of nsp7 and nsp8 confers processivity to nsp12 (Subissi et al. 2014).
The rep proteases that are essential for viral polyprotein processing by the coronaviruses and enteroviruses exhibit a strong preference for substrates containing Gln at P1 position, and share an active-site conformation that engages the substrate's P1 residue. Compound 11r and compound 13b are peptidomimetic α-ketoamides that function as high-affinity non-cleavable substrate analogues and thus exhibit antiviral activity against dimeric 3C-like proteinases (C3Lp dimer) of coronaviruses and enteroviruses (Chen et al. 2005, Zhang et al. 2020). The clinical safety and efficacy of α-ketoamides in Covid-19 are under investigation.
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
To process the nsp3/4 cleavage site, PL-PRO and, presumably, nsp3-4 need to be glycosylated and localized to a membrane (Harcourt et al, 2004).
Virally encoded RNA-dependent RNA polymerase (nsp12, also known as RdRP) is the key component of the replication transcription complex (RTC). SARS-CoV-2-derived nsp12, in complex with nsp7 and nsp8, was shown to have RNA polymerization activity on a poly-U template (Yin et al. 2020). Details of SARS-CoV-2 replication have not yet been elucidated and are inferred from SARS-CoV-1. As SARS-CoV-2 and SARS-CoV-1 are plus strand RNA viruses, nsp12 first synthesizes the complementary minus RNA strand. The purified SARS-CoV-1 nsp12 shows both primer dependent and primer-independent RNA synthesis activities using homopolymeric RNA templates. The catalytic activity of nsp12 is strictly dependent on manganese ions (Mn2+) and primers when the template is a viral-genome-derived RNA representing part of the 3’-UTR of the plus strand with a polyA tail. A 36 nucleotide sequence from the 3’-UTR, predicted to form a stable stem-loop structure, seems to be the minimal cis-acting RNA element required for nsp12 to initiate RNA synthesis (Ahn et al. 2012). The complex of nsp7 and nsp8 confers processivity to nsp12 (Subissi et al. 2014).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Part of nsp4 protein becomes N-glycosylated and gets recruited to the replication complexes in infected cells (Oostra et al, 2007).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Human glycogen synthase kinases GSK3A and GSK3B phosphorylate SARS-CoV-1 coronavirus N (nucleocapsid) protein on multiple serine and threonine residues. GSK3-mediated phosphorylation of the N protein is needed for efficient replication of viral genomic RNA (Wu et al. 2009).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Glycosylated nsp3 (papain-like protease) cleaves the N-proximal polyprotein regions at three sites (Thiel et al, 2003; Harcourt et al, 2004).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
The SARS-CoV nsp3 was shown to bind ORF7a and Nsp6 by using proteomics analysis (Neuman et al, 2008)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
nsp14 acts as 3'-5' exonuclease (Minskaia et al. 2006, Chen et al. 2007) that preferentially excises mismatched nucleotides from double stranded RNA (Minskaia et al. 2006, Bouvet et al. 2012). Binding to nsp10 increases the exonuclease activity of nsp14 (Bouvet et al. 2012, Subissi et al. 2014, Bouvet et al. 2014). nsp14 increases the fidelity of human SARS coronavirus 1 (SARS-CoV-1) replication by the nsp12 RNA-dependent RNA polymerase by 21-fold (Eckerle et al. 2010).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Interaction of the ribonucleocapsid and the structural proteins of SARS-COV-2 in the ERGIC membrane is presumed to promote the formation of virions by budding into to the ERGIC lumen, as is the case for other coronaviruses (reviewed in Masters, 2006). Coronavirus membrane curvature is driven by M lattice formation, interaction with the nucleocapsid and by E protein (de Haan et al, 1998; de Haan et al, 2000; Hsieh et al, 2005; Hsieh et al, 2008; reviewed in Masters, 2006; Ujike and Taguchi, 2015; Schoeman and Fielding 2019).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Similar to other coronaviruses, SARS-CoV-2 virions are released from the host cell by exocytosis in smooth-walled vesicles (reviewed in Ujike and Taguchi, 2015; Masters, 2006; Fung and Liu, 2019). The nature and details of this export remain to be elucidated.
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Transmembrane protease serine 2 (TMPRSS2), associated with the plasma membrane of the host cell, mediates the hydrolytic cleavage of SARS-CoV-2 Spike (S) protein component of the viral membrane-associate S3:M:E:encapsidated SARS coronavirus genomic RNA: 7a:O-glycosyl 3a tetramer complex associated with ACE2 (Matsuyama et al. 2010; Glowacka et al. 2011; Shulla et al. 2011).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Protein 3a can form homodimers and tetramers. The homotetramer shows typical patterns of ion channels. Transport of potassium ions through this channel is effective (Lu et al, 2006). Potassium efflux by protein 3a is important for 3a-induced NLRP3 inflammasome activation (Chen et al, 2019)
Remdesivir (GS-5734) is an investigational nucleotide analogue drug that was developed for its broad spectrum antiviral potential against Ebola and Marburg virus activity (Siegel et al. 2017). It targets and inhibitis viral RNA-dependent RNA polymerase (nsp12, RdRP), the key component of the replication transcription complex (RTC) (Agostini et al. 2018, Brown et al. 2019, Gordon et al. 2020). Remdesivir is being investigated for potential antiviral activity against SARS-CoV-2 by targeting viral replication (Agostini et al. 2018). Gordon et al. demonstrate remdesivir possesses broad antiviral activity against RNA viruses, including SARS-CoV, MERS-CoV and SARS-CoV-2 in-vitro (Gordon et al. 2020b). It could prevent asymptomatic, mild or moderate Covid-19 cases from progressing to severe disease (clinical trials NCT04252664, NCT04257656) but results so far in infected people have been mixed.
EIDD-2801, is an isopropylester prodrug of the ribonucleoside analogue N4-hydroxycytidine (NHC, EIDD-1931) that shows broad spectrum antiviral activity against various RNA viruses including Ebola, Influenza and CoV (Toots et al. 2019). NHC acts as a competitive alternative substrate for virally encoded RNA-dependent RNA polymerases. NHC was shown to inhibit multiple genetically-distinct Bat-CoV viruses in human primary epithelial cells without affecting cell viability. Prophylactic/therapeutic oral administration of NHC reduced lung titers and prevented acute lung failure in C57B/6 mice infected with CoV. The potency of NHC against multiple coronaviruses, its therapeutic efficacy, and oral bioavailability in vivo, all highlight its potential as an effective antiviral against SARS-CoV-2 and other future zoonotic coronaviruses (Sheahan et al. 2020).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Human zinc finger CCHC-type and RNA-binding motif-containing protein 1 (ZCRB1, also known as MADP1) binds to the 5'UTR of the plus strand genomic RNA of the SARS-CoV-1, as well as other coronaviruses, infectious bronchitis virus (IBV) and human coronavirus OC43. ZCRB1 normally localizes to the nucleus, where it is a component of the U12-type spliceosome. Upon infection with a coronavirus, ZCRB1 appears in the cytosol. Binding of ZCRB1 to the 5'UTR stem of coronavirus genomic RNA is thought to be necessary for efficient transcription of viral genes (Tan et al. 2012).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
The SARS-CoV-2 nucleocapsid is released from the host cell endosome into the cytosol. Molecular details of this step are not well worked out. Studies of the infection of the human cultured cells with HCoV-229E coronavirus established a requirement for VCP (transitional endoplasmic reticulum ATPase) protein function for release to occur (Wong et al. 2015). A similar requirement for VCP involvement in SARS-CoV-2 nucleocapsid release is inferred here.
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
A sialyltransferase adds a terminal sialic acid moiety to protein 3a with an O-linked glycosyl side chain. This glycosylated form later is associated with the virion (Oostra et al, 2006)
The function of the non-structural protein nsp16 as a 2'O-methyltransferase that acts in complex with its co-activator nsp10 is conserved in SARS-CoV-2 (Viswanathan et al. 2020).
The genomic and subgenomic mRNAs of SARS-CoV-2 coronavirus, including the plus strand genomic RNA, are presumed to be capped at their 5′ end, based on studies of the mouse hepatitis virus (MHV) (Lai and Stohlman 1981) and the equine torovirus (van Vliet et al. 2002). The non-structural protein 16 (nsp16) acts as a 2'O-methyltransferase that converts coronavirus cap-0 to cap-1, which was first demonstrated with nsp16 cloned from the feline coronavirus (FCV) (Decroly et al. 2008). Cap-0 represents N7-methyl guanosine connected to the 5′ nucleotide through a 5′ to 5′ triphosphate linkage (also known as m7G cap or m7Gppp cap). Cap-1 is generated by an additional methylation on the 2′O position of the initiating nucleotide, and is also known as m7GpppNm. Non-structural protein 10 (nsp10) acts as an activator of nsp16 and is necessary for cap-1 synthesis (Bouvet et al. 2010, Decroly et al. 2011). Coronavirus RNAs with cap-1 are protected from IFIT-mediated interferon response, as IFITs recognize unmethylated 2'-O RNAs. IFITs are interferon-induced proteins with tetratricopeptide repeats that recognize unmethylated 2'-O RNAs and act to inhibit expression of virally encoded mRNAs (Menachery et al. 2014).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
The viral nucleocapsid complex, released into the host cell cytosol, dissociates to release the viral RNA genome (Fung & Liu 2019).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Protein 3a is rapidly internalized into cells by endocytosis. It contains a Yxxφ motif and also diacidic motifs which are typically found in internalized membrane proteins (Tan et al, 2004). The ability to be internalized is necessary for the protein's pro-apoptotic function (Wong et al, 2006; Chan et al, 2009)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Main protease cleaves all cleavage sites of pp1a and ppa1b starting with nsp4/5, thus cleaving itself, and all the cytosolic RTC proteins (Fan et al, 2004)
SARS-CoV-2 transcripts are polyadenylated (Ravindra et al. 2020, Kim et al. 2020), similar to their SARS-CoV-1 counterparts. SARS-CoV-2 subgenomic RNAs (sgRNAs) carry poly(A) tails of the meadian lenght of 47 nucleotides. The poly(A) tails of sgRNAs of SARS-CoV-2 are shorter than the poly(A) tails of the full-length SARS-CoV-2 genomic RNA (Kim et al. 2020).
SARS-CoV-1 plus strand sgRNAs share a 3'UTR with the plus strand genomic RNA, and as this 3'UTR possesses a polyadenylation signal, they undergo polyadenylation by an undetermined viral RNA polymerase, possibly nsp8 or nsp12 (Spagnolo and Hogue 2000, Peng et al. 2016, Tvarogova et al. 2019).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
The genomic and subgenomic mRNAs of SARS-CoV-1 coronavirus, including the plus strand genomic RNA, are presumed to be capped at their 5′ end, based on studies of the mouse hepatitis virus (MHV) (Lai and Stohlman 1981) and the equine torovirus (van Vliet et al. 2002). Non-structural protein 14 (nsp14) acts as an RNA guanine-N7-methyltransferase (N7-MTase) that completes the synthesis of the cap-0 on the SARS-CoV-1 plus strand genomic RNA. Cap-0 represents N7-methyl guanosine connected to the 5′ nucleotide through a 5′ to 5′ triphosphate linkage, and is also known as m7G cap or m7Gppp cap. The N7-MTase domain maps to the carboxy-terminal part of nsp14 (Chen et al. 2009). Cap-0 formation requires three sequential reactions catalyzed by RNA triphosphatase (TPase), guanylyltransferase (GTase), and N7-MTase. There is no evidence that nsp14 possesses TPase and GTase activities, and no other SARS-CoV-1 proteins with these activities have been identified, so the identities of the enzymes that mediate these required steps remain unknown. Based on the study of the human coronavirus 229E, non-structural protein 13 (nsp13) may have a TPase activity in addition to its established helicase activity (Ivanov and Ziebuhr 2004).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Nucleoprotein (N) is ADP-ribosylated. The modification is maintained both in the cell and in virions (Grunewald et al, 2018). Members of the protein mono-ADP-ribosyltransferase (PARP) enzyme family are thought to catalyze this reaction (Fehr et al. 2020)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
Trimmed palmitoylated Spike protein trimers become associated with the ERGIC (ER-Golgi Intermediate Compartment) (Fung & Liu 2019)
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
A minor proportion of the E protein is modified by N-linked glycosylation. This variant appears to be more likely to form multimers, and it shows a different membrane topology than the main variant (Yuan et al, 2006).
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
In the presence of functional nsp14, which acts as a 3'-to-5' exonuclease, the mutation rate during human SARS coronavirus 1 (SARS-CoV-1) replication is 9 x 10^-7 (9E-7) per nucleotide per replication cycle or 2.2 x 10^-5 (2.2E-5) non-redundant substitutions per nucleotide, which translates into 2-3 nucleotide substitutions for each replicated SARS-CoV-1 genome. When nsp14 is defective, the mutation rate during SARS-CoV-1 replication increases to 1.2 x 10^-5 (1.2E-5) mutations per nucleotide per replication cycle or 3.34 x 10^-4 (3.34E-4) non-redundant substitutions per nucleotide, which translates into 12-23 nucleotide substitutions for each replicated SARS-CoV-1 genome (Eckerle et al. 2010). Here the process is annotated in two steps, nsp12- mediated misincorporation of a base (this reaction) and nsp14-mediated detection and removal of that base (next reaction).
Glycosyltransferases in the endoplasmatic reticulum are responsible for the attachment of numerous high-mannose N-glycans on the SARS-CoV-2 spike protein. After virion assembly and release these glycosidations are required for fusion with host cells (Watanabe et al, 2020, Breuer et al, 2001).
Except for position 234, all Man(9)-N-glycan modifications on SARS-Cov-2 Spike are further trimmed to yield high-mannose N-glycan groups containing 5-8 mannose sugars. This reaction is mainly catalyzed by human MAN1B1 glycosdase (Watanabe et al, 2020; Avezov et al, 2008).
Nearly all di-antennary N-glycan sidechains of Spike get further extended to tri-antennary configuration by addition of fucose and further N-acetylglucosamine moieties. Asn-1158 is probably an exception, it stays di-antennary (Watanabe et al, 2020).
Based on SARS-CoV-1 experiments, SARS-CoV-2 virions attached to the host cell surface via a complex involving viral spike (S) protein and host angiotensin-converting enzyme 2 (ACE2) are inferred to undergo endocytosis. In the case of SARS-CoV-1, studies with pseudoviruses have established that S protein is necessary and sufficient for mediating viral attachment and entry. Inhibition of this SARS-CoV-1 S protein-mediated transduction by two different classes of lysosomotropic agents in multiple cell lines strongly suggests that acidification of endosomes is needed for viral entry (Hofmann et al. 2004; Simmons et al. 2004; Yang et al. 2004). The roles of S protein in viral binding to the host cell membrane and fusion of viral and host cell membranes and thus the central role of S protein in determining the host range and tissue tropisms of the virus are reviewed by Belouzard et al. (2012).
This COVID‑19 event has been created by a combination of computational inference from SARS-CoV-1 data (https://reactome.org/documentation/inferred-events) and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
The replicase polyprotein 1a of the human severe acute respiratory syndrome coronavirus (SARS-CoV) is post-translationally cleaved by virally encoded proteases to generate non-structural proteins (nsps). Viral nsps induce the formation of ER-bound double membrane vesicles (DMV) in host cells post infection. These DMVs are decorated with microtubule-associated proteins 1A/1B light chain 3B (MAP1LC3B) proteins that are involved in autophagosome formation. However, there is no evidence that DMVs are recruited to the autophagy machinery. Immunofluorescence studies show that nsp6 (Cottam E M. et al 2011) and nsp8 (Prentice E. et al 2004) colocalizes with MAP1LC3B suggesting a binding event. In some cell types, expression of sars9b (9b) triggers the formation of autophagosomes and underlying molecular mechanisms are unclear (Shi C S. et al 2014). Studies also show that sars8b (8b) can trigger cellular stress, which results in a calcineurin dependent Transcription Factor EB (TFEB) activation and its target genes. This leads to an increase in autophagic flux (Shi C S. et al 2019).
SARS-CoV-2 virions attached to the host cell surface via a complex involving viral spike (S) protein and host angiotensin-converting enzyme 2 (ACE2) are inferred to undergo endocytosis. In the case of SARE-CoV-2 it is apparent that the spike protein undergoes cleavage in some cases and fuses directly with the plasma membrane. This cleavage is induced by FURIN or TMPRSS.
The cellular protease furin cleaves the spike protein at the S1/S2 site and that cleavage is essential for S-protein-mediated cell-cell fusion and entry into human lung cells (Hoffman et al. 2020).
Transmembrane protease serine 2 (TMPRSS2), associated with the plasma membrane of the host cell, mediates the hydrolytic cleavage of SARS-CoV-2 Spike (S) protein component of the viral membrane-associate S3:M:E:encapsidated SARS coronavirus genomic RNA: 7a:O-glycosyl 3a tetramer complex associated with ACE2 (Matsuyama et al. 2010; Glowacka et al. 2011; Shulla et al. 2011).
The uncoated virion nucleocapsid is released directly into the cytoplasm.
This COVID-19 event has been created by a combination of computational inference (see https://reactome.org/documentation/inferred-events) from SARS-CoV-1 data and manual curation, as described in the summation for the overall SARS-CoV-2 infection pathway.
the cellular protease furin cleaves the spike protein at the S1/S2 site and that cleavage is essential for S-protein-mediated cell-cell fusion and entry into human lung cells (Hoffman et al. 2020). Furin is associated with the plasma membrane of the host cell, mediates the hydrolytic cleavage of SARS-CoV-2 Spike (S) protein component of the viral membrane-associate S3:M:E:encapsidated SARS coronavirus genomic RNA: 7a:O-glycosyl 3a tetramer complex associated with ACE2.
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lattice:E
protein:encapsidated SARS-CoV-2 genomic RNA7a:O-glycosyl 3a
tetramerSARS-CoV-2 genomic RNA: 7a:O-glycosyl 3a
tetramer:glycosylated-ACE2SARS-CoV-2 genomic RNA: 7a:O-glycosyl 3a
tetramer:glycosylated-ACE2SARS-CoV-2 genomic RNA: 7a:O-glycosyl
3a tetramerSARS-CoV-2 genomic RNA:7a:O-glycosyl
3a tetramerSARS-CoV-2 genomic RNA:O-glycosyl 3a
tetramergRNA with secondary
structure:RTCgRNA:RTC:nascent RNA minus strand with mismatched
nucleotidegRNA:RTC:RNA primer:RTC
inhibitorsgRNA:RTC:nascent RNA minus strand:RTC
inhibitorsgRNA:RTC:nascent
RNA minus strandcomplement (minus strand):RTC:RTC
inhibitorscomplement (minus
strand):RTCRNA complement
(minus strand)strand subgenomic
mRNAsstrand subgenomic
mRNAsdimer:SARS-CoV-2
genomic RNAN-glycan-PALM-Spike
trimerSARS-CoV-2 genomic
RNA (plus strand)SARS-CoV-2 genomic
RNAN-glycan folded
SpikeN-glycan unfolded
SpikeN-glycan-PALM-Spike
trimerN-glycan-PALM-Spike
trimerSARS-CoV-2 genomic
RNA (plus strand)SARS-CoV-2 genomic RNA complement
(minus strand)polyadenylated SARS-CoV-2 subgenomic mRNAs
(plus strand)polyadenylated SARS-CoV-2 genomic
RNA (plus strand)plus strand
subgenomic mRNAsSARS-CoV-2 genomic
RNA (plus strand)SARS-CoV-2 genomic RNA complement
(minus strand)N-glycan-PALM-Spike
trimerAnnotated Interactions
lattice:E
protein:encapsidated SARS-CoV-2 genomic RNAlattice:E
protein:encapsidated SARS-CoV-2 genomic RNAOtamixaban (FXV673), an anticoagulant, is a potent and selective direct inhibitor of coagulation factor Xa. Virtual docking studies suggest that otamixaban may bind to the serine protease TMPRSS2 (Rensi et al. 2020, preprint). Inhibition of TMPRSS2 is being examined for antiviral activity but its inhibitory potential and/or antiviral activity have not yet been determined so it is annotated here as a candidate drug. I-432 is another inhibitor of TMPRSS2 under investigation for anti viral potential (Pászti-Gere et al. 2016).
Teicoplanin is thought to inhibit the low pH cleavage of the viral spike protein by CTSL in late endosomes thereby preventing the release of genomic viral RNA and the continuation of virus replication cycle (Baron et al. 2020). The target sequence that serve as the cleavage site for CTSL is conserved in the SARS-CoV-2 spike protein (Zhou et al. 2020 [preprint]). Further investigation is required to determine the therapeutic potential of teicoplanin in COVID-19 patients.
Relacatib is an investigational drug trialed for the treatment of osteoporosis (Duong et al. 2016). It is a potent CTSK inhibitor but also shows activity against CTSL (Kumar et al. 2007) so could potentially be investigated for Covid-19 patients. The antileprotic drug clofazimine and the antituberculous drugs rifampicin and isoniazid have been shown to inhibit cathepsins B, H and L from purified goat and bovine brains (Kamboj et al. 2003).
It is generally believed that inhibition of ER glucosidase I and/or II prevents the removal of the terminal glucose moieties on N-linked glycans and results in misfolding and retention of glycoproteins in the ER and ultimate degradation via the ER-associated degradation (ERAD) pathway (Simsek et al. 2005, Alonzi et al. 2013). As a consequence of the abnormal trafficking and degradation of viral glycoproteins, virion assembly and secretion are inhibited (Chang et al. 2009, Taylor et al. 1998).
Long-term suppression of ER glucosidases I and/or II with more potent inhibitors may cause significant side effects, particularly in nerve and immune systems (Sadat et al. 2014).
Li is a selective ATP competitive inhibitor of GSK-3 (Ryves and Harwood, 2001). Lithium carbonate has been and continously is in clinical trials with bipolar disorder patients (Moore et al, 2009). LY2090314 has been in clinical trials for metastatic pancreatic cancer and acute leukemia ([NCT01632306], [NCT01287520], [NCT01214603]). Clinical trials of GSKi for Alzheimer's disease were unsuccessful.
The use of GSKi remains controversial because of their possibly oncogenic properties. Evaluation of GSKi in clinical trials has been hampered by the fear that inhibition of GSK-3 may stimulate or aid in malignant transformation as GSK-3 can phosphorylate pro-oncogenic factors such as beta-catenin, c-Jun and c-Myc which targets them for degradation (Patel & Woodgett, 2008). However, no studies have been reported suggesting that treatment of mice with GSKi resulted in an increase in cancer incidence. In fact, many patients with bi-polar disorder have been treated with lithium for prolonged periods of time. There does not appear to be any evidence that these patients have increased incidences of cancer (McCubrey et al, 2014).
The GSKi kenpaullone and lithium chloride were found to reduce viral Nucleoprotein phosphorylation in the severe acute respiratory syndrome CoV-infected VeroE6 cells and decrease the viral titer and cytopathic symptoms. Effect of GSK-3 inhibition were reproduced in another coronavirus, the neurotropic JHM strain of mouse hepatitis virus (Wu et al, 2009).
nsp13 of SARS-CoV-1 is an ATP-dependent helicase that functions in the 5'-3' direction to unwind double stranded RNAs that have a 5' single strand overhang at least 20 nucleotides long. nsp13 can also act on double strand DNA in vitro, but dsRNA is thought to be its physiological substrate. The catalytic activity of SARS-CoV-1 nsp13 is increased in the presence of nsp12, the viral RNA-dependent RNA polymerase. nsp13 is needed for the replication of SARS-CoV-1 and is thought to act by melting secondary structures in the genomic RNA template during replication, and also to be involved in unwinding of RNA duplexes during transcription of viral genes. nsp13 is a promising target for experimental anti-SARS-CoV-1 drugs (Tanner et al. 2003, Ivanov et al. 2004, Bernini et al. 2006, Chen et al. 2009, Lee et al. 2010, Adedeji et al. 2012).
In about 15% of translation attempts of the genomic viral mRNA1 in the host cell cytosol, a -1 frameshift happens after the nsp10 gene that leads to translation of the full pp1ab polyprotein (Baranov et al, 2005).
nsp8 functions as an RNA-dependent RNA polymerase (RdRp) that serves as the primase for nsp12, the main RdRp of the SARS coronavirus 1 (SARS-CoV-1) (Imbert et al. 2006), as it is capable of de novo RNA synthesis (te Velthuis et al. 2011). nsp8 synthesizes short oligonucleotides (less than 6 bases long) using genomic RNA as a template. nsp8 requires at least one cytidine residue in the template sequence for its activity. Activity is dependent on manganese ions (Imbert et al. 2006). nsp8 can also extend primers but is 20-fold less efficient than nsp12 (te Velthuis et al. 2011).
Based on studies in other coronaviruses, the final SARS-COV-2 ribonucleoprotein complex is predicted to be a hollow helical structure with an approximate diameter of 9-16nm, with the C-terminal domain of N protein forming the inner core and the N-terminal domain forming the outer surface (Neuman et al, 2006; Chen et al, 2007). Oliomerization of the N protein capsid coat is likely nucleated through both protein-RNA and protein-protein interactions by the first few N-protein dimers on the genomic RNA (Saikatendu et al, 2007; Chang et al, 2013; reviewed in Chang et al, 2014). Each N dimer may make contact with up to 7 bases of the RNA (reviewed in Chang et al, 2014).
Within the host cell endocytic vesicle, SARS-CoV-2 Spike (S) protein is cleaved between residues 797 and 798 by cathepsin L1 (CTSL) (Huang et al. 2006). The roles of S protein in viral binding to the host cell membrane and fusion of viral and host cell membranes and thus the central role of S protein in determining the host range and tissue tropisms of the virus are reviewed by Belouzard et al. (2012).
The majority of nucleoprotein is serine-phosphorylated in the cytosol and, possibly, in the nucleus where it gets immediately transported to the cytosol. Phosphorylation is catalyzed by glycogen synthase kinase 3 (GSK3) and several other host cell kinases (Surjit et al, 2005; Wu et al. 2009)
Both a predicted beta-hairpin motif and the N-terminal part of protein E are sufficient for its localization to the Golgi membrane. Although porin activity has been shown for protein E it cannot be detected in the plasma membrane of infected cells (Liao et al, 2006; Cohen et al, 2011; Nieto-Torres, 2011).
Non-structural protein 14 (nsp14) of the human SARS coronavirus is a bifunctional enzyme bearing 3'-5 exoribonuclease activity involved in replication fidelity and RNA cap N7-guanine methyltransferase activity involved in 5'-RNA capping. nsp14 binds to the minimal replication and transcription complex (RTC), composed of nsp7, nsp8, and nsp12, by directly binding to nsp12 (the main RNA-dependent RNA polymerase). Binding of nsp14 does not affect the processivity of the RTC (Minskaia et al. 2006, Subissi et al. 2014).
Membranous structures containing protein 3a are being shedded from the cell membrane (Huang et al, 2006)
nsp3 and nsp4 alone caused considerable ER membrane deformation, producing a perinuclear double-walled maze-like body (MLB), and the nsp3–nsp4 interaction was shown to be absolutely necessary for such membrane rearrangement. Further necessary factors are nsp6 and unidentified host factors (Angelini et al 2013, Sakai et al 2017). nsp6 by itself can form Atg5 and LC3II-positive vesicles classically observed in autophagy (Cottam et al, 2014). However, in mouse hepatitis virus (MHV) infections, EDEM1 and OS9 of the ER-associated degradation system have been shown to be necessary co-factors (Reggiori et al, 2010)
To process the nsp3/4 cleavage site, PL-PRO and, presumably, nsp3-4 need to be glycosylated and localized to a membrane (Harcourt et al, 2004)
In most translation attempts the genomic viral mRNA1 in the cytosol is translated to a shortened polyprotein, pp1a, that does not contain genome replication enzymes (Baranov et al, 2005).
Calnexin transiently binds the unfolded spike protein and prevents its aggregation and premature degradation, ensuring its correct folding (Fukushi et al, 2012).
The papain-like protease domain of the nsp1-4 fragment alone is sufficient for processing the nsp1/2 and nsp2/3 cleavage sites (Harcourt et al, 2004).
Two of the four cysteine-rich clusters of the viral spike protein are modified by palmitoylation. This is required for the protein's partitioning into detergent-resistant membranes and for cell–cell fusion. In general, palmitoylation is usually non-enzymatic (Petit et al, 2007; McBride and Machamer, 2010; Veit, 2012).
SARS-CoV-1 encodes eight subgenomic RNAs, mRNA2 to mRNA9. mRNA1 corresponds to the genomic RNA. The 5' and 3' ends of subgenomic RNAs are identical, in accordance with the template switch model of coronavirus RNA transcription (Snijder et al. 2003, Thiel et al. 2003, Yount et al. 2003). Therefore, consistent with this and the studies of the murine hepatitis virus (MHV), which is closely related to SARS-CoV-1, genomic positive strand RNA is first transcribed into negative sense (minus strand) subgenomic mRNAs, that subsequently serve as templates for the synthesis of positive strand subgenomic mRNAs. Negative-sense virus RNAs are present in much smaller amounts than positive-sense RNAs (Irigoyen et al. 2016). Each subgenomic RNA contains a leader transcription regulatory sequence (leader TRS) that is identical to the leader of the genome, appended via polymerase “jumping� during negative strand synthesis to the body transcription regulatory sequence (body TRS), a short, AU-rich motif of about 10 nucleotides found upstream of each ORF that is destined to become 5' proximal in one of the subgenome-length mRNAs. The 3' and 5'UTRs may interact through RNA–RNA and/or RNA–protein plus protein–protein interactions to promote circularization of the coronavirus genome, placing the elongating minus strand in a favorable topology for leader-body joining. The host protein PABP was found to bind to the coronavirus 3' poly(A) tail and to interact with the host protein eIF-4G, a component of the three-subunit complex that binds to mRNA cap structures, which may promote the circularization of the coronavirus genome. Two viral proteins that bind to the coronavirus 5'UTR, the N protein and nsp1, may play a role in template switching. The poly(A) tail is necessary for the initiation of minus-strand RNA synthesis at the 3' end of genomic RNA. For review, please refer to Sawicki et al. 2007 and Yang and Leibowitz 2015.
A certain part of the nucleoprotein can be found in the nucleolus. This localisation seems to depend on the protein's sumoylation (Li et al, 2005)
Protein M accumulates in the Golgi complex and recruits Spike protein to the sites of virus assembly and budding in the ERGIC (Voss et al, 2009).
Significant amounts of the unphosphorylated N protein are associated with the cell membrane (Surjit et al, 2005)
The crucial step of autocleavage of pp1a involves the formation of an "intermediate" pp1a dimer which has weak protease activity. This "embedded" 3CLp liberates itself by cleaving the ends off its monomer in trans. Only after that occurs does the cleaved 3CLp form a dimer, the most efficient form of the enzyme (Hsu et al, 2005; Chen et al, 2010; Muramatsu et al, 2016)
To process the nsp3/4 cleavage site, PL-PRO and, presumably, nsp3-4 need to be glycosylated and localized to a membrane (Harcourt et al, 2004)
Human SARS coronavirus 1 (SARS-CoV-1) non-structural protein 15 (nsp15) contains the LXCXE/D motif characteristic of proteins that bind to the retinoblastoma protein RB1. Binding to human RB1 increases the endonuclease activity of nsp15 but is not required for it. RB1 bound to nsp15 is retained in the cytosol. Interaction of nsp15 with RB1 likely affects the cell cycle of infected cells and probably modulates cytotoxicity of SARS-CoV-1 (Bhardwaj et al. 2012).
Viral protein 3a translocates from the cytosol to the ERGIC (endoplasmic reticulum Golgi intermediate compartment) (Oostra et al. 2006)
Viral E protein is modified by palmitoylation at all three cysteine residues. In general, palmitoylation is usually non-enzymatic (Liao et al, 2006, Veit, 2012).
nsp13, the helicase of the human SARS coronavirus 1 (SARS-CoV-1) binds to DDX5, a host protein implicated in transcription, pre-mRNA processing, RNA degradation, RNA export, ribosome assembly and translation. DDX5 knockdown inhibits viral replication (Chen et al. 2009).
EIDD-2801, is an isopropylester prodrug of the ribonucleoside analogue N4-hydroxycytidine (NHC, EIDD-1931) that shows broad spectrum antiviral activity against various RNA viruses including Ebola, Influenza and CoV (Toots et al. 2019). NHC acts as a competitive alternative substrate for virally encoded RNA-dependent RNA polymerases. NHC was shown to inhibit multiple genetically-distinct Bat-CoV viruses in human primary epithelial cells without affecting cell viability. Prophylactic/therapeutic oral administration of NHC reduced lung titers and prevented acute lung failure in C57B/6 mice infected with CoV. The potency of NHC against multiple coronaviruses, its therapeutic efficacy, and oral bioavailability in vivo, all highlight its potential as an effective antiviral against SARS-CoV-2 in Covid-19 infections and other future zoonotic coronaviruses (Sheahan et al. 2020).
A sialyltransferase adds a terminal sialic acid moiety to protein 3a with an O-linked glycosyl side chain. This glycosylated form later is associated with the virion (Oostra et al, 2006)
Main protease cleaves all cleavage sites of pp1a and ppa1b starting with nsp4/5, thus cleaving itself, and all the cytosolic RTC proteins (Fan et al, 2004)
The ribonucleoprotein complex is recruited to the assembling virion through interactions with the M C-terminal tail, and appears to be independent of viral RNA (Hsieh et al, 2008; Hatakeyama et al, 2008; He et al, 2004; Luo et al, 2006; reviewed in Ujike and Taguchi, 2015). How much of this is fully conserved in SARS-COV-2 remains to be experimentally verified.
In SARS-CoV-1, nsp16 was identified as an AdoMet-dependent (nucleoside-2'O)-methyltransferase involved in capping of viral RNAs. nsp16 binds to nsp10, which serves as a cofactor for nsp16 (Bouvet et al. 2010, Lugari et al. 2010). Nsp16 alone is unstable and exhibits 2'-O-methyltransferase activity only in complex with nsp10 (Debarnot et al, 2011; Decroly et al, 2011). nsp10-mediated activation of nsp16 catalytic activity is conserved in all coronaviruses (Wang et al. 2015). The same binding surface of nsp10 interacts with nsp14 and nsp16, suggesting that binding of nsp14 and nsp16 to nsp10 is mutually exclusive. However, as nsp10 is produced in a higher number of copies than nsp14 and nsp16, and as nsp14 and nsp16 act coordinately in RNA capping, it is most likely that nsp14:nsp10 and nsp16:nsp10 complexes co-exist within the viral replication-transcription complex (RTC) (Bouvet et al. 2012, Bouvet et al. 2014). One structural study reported that nsp10 forms dodecamers (Su et al. 2006), which would potentially allow simultaneous binding of nsp14 and nsp16 to nsp10 homomeric complexes, but it is not certain if such homomeric complexes of nsp10 exist in vivo, and if the structure of the nsp10 dodecamer would be permissive for nsp16 binding (Chen et al. 2011). nsp10 contains two zinc fingers which are thought to be involved in RNA binding (Su et al. 2006, Joseph et al. 2006).
Protein E forms a pentamer of monomers without disulfide bonds (Parthasarathy et al, 2012)
Based on studies in SARS-CoV-1, the RTC binds to the 3' end of the viral plus strand genomic RNA to initiate synthesis of the complementary minus strand. A 36 nucleotide sequence from the 3’-UTR of the plus strand, predicted to form a stable stem-loop structure, seems to be the minimal cis-acting RNA element required for the viral RNA-directed RNA polymerase (nsp12) to initiate RNA synthesis. The polyA tail also seems to play a role in the initiation of replication of viral genomic RNA (Ahn et al. 2012).
Binding of the genomic RNA to one or a small number of N-N dimers may be the initiating event in nucleocapsid formation. Both the NTD and the CTD of N have been shown to have RNA-binding activity (Huang et al, 2004a; Huang et al, 2004b; Chen et al, 2007; Chang et al, 2009; Takeda et al, 2008), and the IDRs seem likely to also contribute (Chang et al, 2009). These initial binding events may nucleate nucleocapsid formation through further recruitment of N protein dimers (reviewed in Chang et al, 2014).
N protein is sumoylated at a lysine residue. Abolition of sumoylation of nucleoprotein significantly decreases homo-oligomerisation of the protein (Li et al, 2005)
The genomic and subgenomic (sg) mRNAs of SARS-CoV-1 coronavirus are presumed to be capped at their 5′ end, based on studies of the mouse hepatitis virus (MHV) (Lai and Stohlman 1981) and the equine torovirus (van Vliet et al. 2002). Non-structural protein 14 (nsp14) acts as an RNA guanine-N7-methyltransferase (N7-MTase) that completes the synthesis of the cap-0 on SARS-CoV-1 mRNAs. The cap-0 represents N7-methyl guanosine connected to the 5′ nucleotide through a 5′ to 5′ triphosphate linkage, and is also known as m7G cap or m7Gppp cap. The N7-MTase domain maps to the carboxy-terminal part of nsp14 (Chen et al. 2009). Cap-0 formation requires three sequential reactions catalyzed by RNA triphosphatase (TPase), guanylyltransferase (GTase), and N7-MTase. There is no evidence that nsp14 possesses TPase and GTase activities, and no other SARS-CoV-1 proteins with these activities have been identified, so the identities of the enzymes that mediate these required steps remain unknown. Based on the study of the human coronavirus 229E, non-structural protein 13 (nsp13) may have a TPase activity in addition to its established helicase activity (Ivanov and Ziebuhr 2004).
Despite its importance, expression of M alone is not sufficient to drive formation of a mature virus (reviewed in Masters, 2006). Protein-protein interactions between M and S, N and E, among other components, are required for assembly of a mature virus and for membrane curvature. Many studies have examined the minimal system required for release of viral like particles (VLPs) with sometimes contradictory results, but interactions between M, N and E are sufficient to promote release of significant numbers of VLPs (Ho et al, 2004; Huang et al, 2004; Mortola and Roy, 2004; Hsieh et al, 2005; Siu et al, 2008; Hatakeyama et al, 2008; Tseng et al, 2013; reviewed in Masters, 2006)
The genomic and subgenomic mRNAs of SARS-CoV-1 coronavirus, including the minus strand genomic RNA complement, are presumed to be capped at their 5′ end, based on studies of the mouse hepatitis virus (MHV) (Lai and Stohlman 1981) and the equine torovirus (van Vliet et al. 2002). The non-structural protein 14 (nsp14) acts as an RNA guanine-N7-methyltransferase (N7-MTase) that completes the synthesis of the cap-0 on SARS-CoV-1 minus strand genomic RNA. The cap-0 represents N7-methyl guanosine connected to the 5′ nucleotide through a 5′ to 5′ triphosphate linkage, and is also known as m7G cap or m7Gppp cap. The N7-MTase domain maps to the carboxy-terminal part of nsp14 (Chen et al. 2009). Cap-0 formation requires three sequential reactions catalyzed by RNA triphosphatase (TPase), guanylyltransferase (GTase), and N7-MTase. There is no evidence that nsp14 possesses TPase and GTase activities, and no other SARS-CoV-1 proteins with these activities have been identified, so the identities of the enzymes that mediate these required steps remain unknown. Based on the study of the human coronavirus 229E, non-structural protein 13 (nsp13) may have a TPase activity in addition to its established helicase activity (Ivanov and Ziebuhr 2004).
After synthesizing the complementary minus RNA of the plus strand viral genomic RNA, SARS-CoV-1 replication-transcription complex (RTC) associates with the minus strand to initiate plus strand synthesis and to initiate transcription of subgenomic (sg) mRNAs (Ahn et al. 2012).
The subgenomic mRNAs of SARS-CoV-2 coronavirus are presumed to be capped at their 5′ ends, based on studies of the mouse hepatitis virus (MHV) (Lai and Stohlman 1981) and the equine torovirus (van Vliet et al. 2002). The non-structural protein 16 (nsp16) acts as a 2'O-methyltransferase that converts coronavirus cap-0 to cap-1, which was first demonstrated with nsp16 cloned from the feline coronavirus (FCV) (Decroly et al. 2008). Cap-0 represents N7-methyl guanosine connected to the 5′ nucleotide through a 5′ to 5′ triphosphate linkage (also known as m7G cap or m7Gppp cap). Cap-1 is generated by an additional methylation on the 2′O position of the initiating nucleotide, and is also known as m7GpppNm. The non-structural protein 10 (nsp10) acts as an activator of nsp16 and is necessary for cap-1 synthesis (Bouvet et al. 2010, Decroly et al. 2011). Coronavirus RNAs with cap-1 are protected from IFIT-mediated interferon response, as IFITs recognize unmethylated 2'-O RNAs. IFITs are interferon-induced proteins with tetratricopeptide repeats that recognize unmethylated 2'-O RNAs and act to inhibit expression of virally encoded mRNAs (Menachery et al. 2014).
SARS-CoV-1 encodes eight subgenomic RNAs, mRNA2 to mRNA9. mRNA1 corresponds to the genomic RNA. mRNA2 encodes the S protein. mRNA3 is bicistronic and encodes proteins 3a and 3b. mRNA4 encodes the E protein. mRNA5 encodes the M protein. mRNA6 encodes the protein 6. mRNA7, mRNA8 and mRNA9 are bicistronic, with mRNA7 encoding proteins 7a and 7b, mRNA8 encoding proteins 8a and 8b, and mRNA 9 encoding proteins 9a and N. The 5' and 3' ends of subgenomic RNAs are identical, in accordance with the template switch model of coronavirus RNA transcription (Snijder et al. 2003, Thiel et al. 2003, Yount et al. 2003). Based on studies of the murine hepatitis virus (MHV), which is closely related to SARS-CoV-1, positive-sense virus mRNAs are present at much higher amounts than negative-sense mRNAs (Irigoyen et al. 2016).
Human von Hippel Lindau (VHL) protein, a tumor suppressor that acts as a component of an E3 ubiquitin ligase complex, interacts with the non-structural protein 16 (nsp16) of the human SARS coronavirus 1 (SARS-CoV-1) and the mouse hepatitis virus, also a coronavirus. VHL negatively regulates SARS-CoV-1 replication, but the exact mechanism is not known (Yu et al. 2015).
The genomic and subgenomic mRNAs of SARS-CoV-2 coronavirus, including the minus strand genomic RNA, are presumed to be capped at their 5′ end, based on studies of the mouse hepatitis virus (MHV) (Lai and Stohlman 1981) and the equine torovirus (van Vliet et al. 2002). Non-structural protein 16 (nsp16) acts as a 2'O-methyltransferase that converts coronavirus cap-0 to cap-1, which was first demonstrated with nsp16 cloned from the feline coronavirus (FCV) (Decroly et al. 2008). Cap-0 represents N7-methyl guanosine connected to the 5′ nucleotide through a 5′ to 5′ triphosphate linkage (also known as m7G cap or m7Gppp cap). Cap-1 is generated by an additional methylation on the 2′O position of the initiating nucleotide, and is also known as m7GpppNm. Non-structural protein 10 (nsp10) acts as an activator of nsp16 and is necessary for cap-1 synthesis (Bouvet et al. 2010, Decroly et al. 2011). Coronavirus RNAs with cap-1 are protected from IFIT-mediated interferon response. IFITs are interferon-induced proteins with tetratricopeptide repeats that recognize unmethylated 2'-O RNAs and act to inhibit expression of virally encoded mRNAs (Menachery et al. 2014).
Protein M is exclusively N-glycosylated at one asparagine by an unknown glycosyltransferase. However, further processing of N-linked glycans is prevented in infected cells. Both the glycosylated and nonglycosylated M is incorporated into the virion. In summary, glycosylation of M is neither a prerequisite for intra-cellular transport nor for recruitment into the virion (Voss et al, 2006).
In addition to the main structural proteins and the nucleocapsid, the mature virion may also contain low proportions of accessory proteins, including protein 3a and 7a (reviewed in McBride and Fielding, 2012). Protein 3a has been shown to interact with E, M, S and protein 7a and is estimated to be present in the virion at 2/3 the molar ratio of E protein (Ito et al, 2005; Shen et al, 2005; Tan et al, 2004). Although 3a tetramers are predicted to act as ion channels in the host plasma membrane, increasing cell permeability, the role of 3a in the mature virion is not clear (Lu et al, 2006; reviewed in McBride and Fielding, 2012). Protein 7a is type 1 transmembrane protein that interacts with M, E, S and protein 3a and may be incorporated into the mature virion. What functional role protein 7a may play in the assembled virion is unclear (Huang et al, 2006; Fielding et al, 2004; Tan et al, 2004).
Viral E protein is ubiquitinated both in vitro and in cells (Alvarez et al, 2011).
Nonstructural protein 15 (nsp15) of the SARS coronavirus (SARS-CoV-1) binds to the replication-transcription complex (RTC) through interaction with nsp8 (Imbert et al. 2008). This interaction appears to be conserved in other coronaviruses, such as mouse hepatitis virus (MHV) (Athmer et al. 2017). nsp15 is an endonuclease characteristic for the order Nidovirales that includes the family Coronaviridae. nsp15 preferentially cleaves 3' of uridines, generating 2'-3' cyclic phosphates after cleavage. nsp15 requires Mn2+ ions for catalytic activity. Functional nsp15 is needed for production of viable virions and for viral transcription (Guarino et al. 2005, Ricagno et al. 2006, Bhardwaj et al. 2006, Joseph et al. 2007, Bhardwaj et al. 2008, Bhardwaj et al. 2012). The biological role of nsp15 has not been elucidated. It may degrade host mRNAs to shut down host translation, but so far no human or viral RNA targets have been identified.
EIDD-2801, is an isopropylester prodrug of the ribonucleoside analogue N4-hydroxycytidine (NHC, EIDD-1931) that shows broad spectrum antiviral activity against various RNA viruses including Ebola, Influenza and CoV (Toots et al. 2019). NHC acts as a competitive alternative substrate for virally encoded RNA-dependent RNA polymerases. NHC was shown to inhibit multiple genetically-distinct Bat-CoV viruses in human primary epithelial cells without affecting cell viability. Prophylactic/therapeutic oral administration of NHC reduced lung titers and prevented acute lung failure in C57B/6 mice infected with CoV. The potency of NHC against multiple coronaviruses, its therapeutic efficacy, and oral bioavailability in vivo, all highlight its potential as an effective antiviral against SARS-CoV-2 and other future zoonotic coronaviruses (Sheahan et al. 2020).
The replication-transcription complex (RTC) completes synthesis of the genomic RNA complement (minus strand). The complex of nsp7 and nsp8 confers processivity to nsp12, the virally encoded RNA-dependent RNA polymerase that replicates the viral genomic RNA, enabling the RTC to complete the RNA synthesis with a very low dissociation rate. nsp7 plays a crucial role in maintaining binding of the RTC to the RNA. nsp14 subunit of the RTC does not affect the processivity (Subissi et al. 2014).
Main protease cleaves all cleavage sites of pp1a and ppa1b starting with nsp4/5, thus cleaving itself, and all the cytosolic RTC proteins (Fan et al, 2004)
Based on studies in other coronaviruses, SARS-COV-2 S trimers are presumed to be recruited to the assembling virion through interaction with M protein (reviewed in Ujike and Taguchi, 2015). Multiple regions of M contribute to the recruitment of S, with a single tyrosine residue in the C-terminal domain of M playing a critical role (McBride et al, 2010a; Hsieh et al, 2008). Interaction with M is aided by a dibasic motif in the C-terminus of S, which promotes retrieval of the spike protein from the cell surface by binding the COPI coat (McBride et al, 2007; Ujike et al, 2016). Palmitoylation of the C-terminus of S appears dispensible for the interaction with M in SARS-COV-1, unlike the case in other coronaviruses; whether this is also true for SARS-COV-2 remains to be determined (Ujike et al, 2012; McBride 2010b; reviewed in Ujike and Taguchi, 2015). Size estimates and modelling suggest the mature virion has approximately 300 S trimers (Neuman et al, 2006; reviewed in Chang et al, 2014).
Protein M accumulates in the Golgi complex and recruits Spike protein to the sites of virus assembly and budding in the ERGIC (Voss et al, 2009).
In the host cell cytosol the pp1a polyprotein spontaneously dimerizes. This temporary dimer has weak protease activity (Chen et al, 2010)
As early as 3 hours post-infection, cytoplasmic accumulations of N are formed in infected cells, they colocalize with viral RNA. From 5 hours post-infection on, N can be detected in the Golgi, the budding site (Stertz et al, 2007)
As it contains cargo sorting motifs in its cytoplasmic domain, protein 3a gets localized by the cell's protein transport system to the cell membrane where it functions as an ion channel (Tan et al, 2004). This ion channel function is necessary for the protein's pro-apoptotic function (Chan et al, 2009)
Some phosphorylated N is found to associate with the cell membrane (Surjit et al, 2005).
On the final assembly, proteomics data suggest that the NAB−βSM−TM1 domains of nsp3 can interact with nsp7 − 8, as well as nsps 12–16, and the domain Y1 plus CoV-Y interacts with nsp9 and nsp12 (Imbert et al., 2008). Also a PL2pro−NAB−βSM−TM1 construct of Nsp3 can bind Nsp4 and Nsp12, while the region from TM1 to the end of Nsp3 only binds Nsp8 (Pan et al., 2008).
SARS-CoV-2 spike (S3a) protein, as a component of the S3:M:E:encapsidated SARS coronavirus genomic RNA: 7a:O-glycosyl 3a tetramer complex, binds to glycosylated angiotensin converting enzyme 2 (ACE2) associated with the human host cell plasma membrane. Structural studies of the interaction between human ACE2 protein and the receptor-binding domain of S3a protein have identified key amino acid residues in both proteins responsible for their high-affinity interaction. These residues may be a key factor determining severity (and possibly human-to-human transmission) of SARS-CoV-2 (Li et al. 2003, 2005). The roles of S protein in viral binding to the host cell membrane and fusion of viral and host cell membranes and thus the central role of S protein in determining the host range and tissue tropisms of the virus are reviewed by Belouzard et al. (2012).
The replicase polyprotein 1a of the human severe acute respiratory syndrome coronavirus is post-translationally cleaved by virally encoded proteases to generate non-structural proteins (nsps). Viral nsps induce the formation of ER-bound double membrane vesicles (DMV) in host cells post infection. These DMVs are decorated with host microtubule-associated proteins 1A/1B light chain 3B (MAP1LC3B) proteins that are involved in autophagosome formation. However, there is no evidence that DMVs are recruited to the autophagy machinery. Immunofluorescence studies show that nsp8 colocalizes with MAP1LC3B suggesting a binding event (Prentice E. et al 2004).
To process the nsp3/4 cleavage site, PL-PRO and, presumably, nsp3-4 need to be glycosylated and localized to a membrane (Harcourt et al, 2004).
Part of nsp4 protein becomes N-glycosylated and gets recruited to the replication complexes in infected cells (Oostra et al, 2007).
Human glycogen synthase kinases GSK3A and GSK3B phosphorylate SARS-CoV-1 coronavirus N (nucleocapsid) protein on multiple serine and threonine residues. GSK3-mediated phosphorylation of the N protein is needed for efficient replication of viral genomic RNA (Wu et al. 2009).
Glycosylated nsp3 (papain-like protease) cleaves the N-proximal polyprotein regions at three sites (Thiel et al, 2003; Harcourt et al, 2004).
The SARS-CoV nsp3 was shown to bind ORF7a and Nsp6 by using proteomics analysis (Neuman et al, 2008)
nsp14 acts as 3'-5' exonuclease (Minskaia et al. 2006, Chen et al. 2007) that preferentially excises mismatched nucleotides from double stranded RNA (Minskaia et al. 2006, Bouvet et al. 2012). Binding to nsp10 increases the exonuclease activity of nsp14 (Bouvet et al. 2012, Subissi et al. 2014, Bouvet et al. 2014). nsp14 increases the fidelity of human SARS coronavirus 1 (SARS-CoV-1) replication by the nsp12 RNA-dependent RNA polymerase by 21-fold (Eckerle et al. 2010).
Interaction of the ribonucleocapsid and the structural proteins of SARS-COV-2 in the ERGIC membrane is presumed to promote the formation of virions by budding into to the ERGIC lumen, as is the case for other coronaviruses (reviewed in Masters, 2006). Coronavirus membrane curvature is driven by M lattice formation, interaction with the nucleocapsid and by E protein (de Haan et al, 1998; de Haan et al, 2000; Hsieh et al, 2005; Hsieh et al, 2008; reviewed in Masters, 2006; Ujike and Taguchi, 2015; Schoeman and Fielding 2019).
Similar to other coronaviruses, SARS-CoV-2 virions are released from the host cell by exocytosis in smooth-walled vesicles (reviewed in Ujike and Taguchi, 2015; Masters, 2006; Fung and Liu, 2019). The nature and details of this export remain to be elucidated.
Transmembrane protease serine 2 (TMPRSS2), associated with the plasma membrane of the host cell, mediates the hydrolytic cleavage of SARS-CoV-2 Spike (S) protein component of the viral membrane-associate S3:M:E:encapsidated SARS coronavirus genomic RNA: 7a:O-glycosyl 3a tetramer complex associated with ACE2 (Matsuyama et al. 2010; Glowacka et al. 2011; Shulla et al. 2011).
Protein 3a can form homodimers and tetramers. The homotetramer shows typical patterns of ion channels. Transport of potassium ions through this channel is effective (Lu et al, 2006). Potassium efflux by protein 3a is important for 3a-induced NLRP3 inflammasome activation (Chen et al, 2019)
EIDD-2801, is an isopropylester prodrug of the ribonucleoside analogue N4-hydroxycytidine (NHC, EIDD-1931) that shows broad spectrum antiviral activity against various RNA viruses including Ebola, Influenza and CoV (Toots et al. 2019). NHC acts as a competitive alternative substrate for virally encoded RNA-dependent RNA polymerases. NHC was shown to inhibit multiple genetically-distinct Bat-CoV viruses in human primary epithelial cells without affecting cell viability. Prophylactic/therapeutic oral administration of NHC reduced lung titers and prevented acute lung failure in C57B/6 mice infected with CoV. The potency of NHC against multiple coronaviruses, its therapeutic efficacy, and oral bioavailability in vivo, all highlight its potential as an effective antiviral against SARS-CoV-2 and other future zoonotic coronaviruses (Sheahan et al. 2020).
Human zinc finger CCHC-type and RNA-binding motif-containing protein 1 (ZCRB1, also known as MADP1) binds to the 5'UTR of the plus strand genomic RNA of the SARS-CoV-1, as well as other coronaviruses, infectious bronchitis virus (IBV) and human coronavirus OC43. ZCRB1 normally localizes to the nucleus, where it is a component of the U12-type spliceosome. Upon infection with a coronavirus, ZCRB1 appears in the cytosol. Binding of ZCRB1 to the 5'UTR stem of coronavirus genomic RNA is thought to be necessary for efficient transcription of viral genes (Tan et al. 2012).
The SARS-CoV-2 nucleocapsid is released from the host cell endosome into the cytosol. Molecular details of this step are not well worked out. Studies of the infection of the human cultured cells with HCoV-229E coronavirus established a requirement for VCP (transitional endoplasmic reticulum ATPase) protein function for release to occur (Wong et al. 2015). A similar requirement for VCP involvement in SARS-CoV-2 nucleocapsid release is inferred here.
A sialyltransferase adds a terminal sialic acid moiety to protein 3a with an O-linked glycosyl side chain. This glycosylated form later is associated with the virion (Oostra et al, 2006)
The genomic and subgenomic mRNAs of SARS-CoV-2 coronavirus, including the plus strand genomic RNA, are presumed to be capped at their 5′ end, based on studies of the mouse hepatitis virus (MHV) (Lai and Stohlman 1981) and the equine torovirus (van Vliet et al. 2002). The non-structural protein 16 (nsp16) acts as a 2'O-methyltransferase that converts coronavirus cap-0 to cap-1, which was first demonstrated with nsp16 cloned from the feline coronavirus (FCV) (Decroly et al. 2008). Cap-0 represents N7-methyl guanosine connected to the 5′ nucleotide through a 5′ to 5′ triphosphate linkage (also known as m7G cap or m7Gppp cap). Cap-1 is generated by an additional methylation on the 2′O position of the initiating nucleotide, and is also known as m7GpppNm. Non-structural protein 10 (nsp10) acts as an activator of nsp16 and is necessary for cap-1 synthesis (Bouvet et al. 2010, Decroly et al. 2011). Coronavirus RNAs with cap-1 are protected from IFIT-mediated interferon response, as IFITs recognize unmethylated 2'-O RNAs. IFITs are interferon-induced proteins with tetratricopeptide repeats that recognize unmethylated 2'-O RNAs and act to inhibit expression of virally encoded mRNAs (Menachery et al. 2014).
The viral nucleocapsid complex, released into the host cell cytosol, dissociates to release the viral RNA genome (Fung & Liu 2019).
Protein 3a is rapidly internalized into cells by endocytosis. It contains a Yxxφ motif and also diacidic motifs which are typically found in internalized membrane proteins (Tan et al, 2004). The ability to be internalized is necessary for the protein's pro-apoptotic function (Wong et al, 2006; Chan et al, 2009)
Main protease cleaves all cleavage sites of pp1a and ppa1b starting with nsp4/5, thus cleaving itself, and all the cytosolic RTC proteins (Fan et al, 2004)
SARS-CoV-1 plus strand sgRNAs share a 3'UTR with the plus strand genomic RNA, and as this 3'UTR possesses a polyadenylation signal, they undergo polyadenylation by an undetermined viral RNA polymerase, possibly nsp8 or nsp12 (Spagnolo and Hogue 2000, Peng et al. 2016, Tvarogova et al. 2019).
The genomic and subgenomic mRNAs of SARS-CoV-1 coronavirus, including the plus strand genomic RNA, are presumed to be capped at their 5′ end, based on studies of the mouse hepatitis virus (MHV) (Lai and Stohlman 1981) and the equine torovirus (van Vliet et al. 2002). Non-structural protein 14 (nsp14) acts as an RNA guanine-N7-methyltransferase (N7-MTase) that completes the synthesis of the cap-0 on the SARS-CoV-1 plus strand genomic RNA. Cap-0 represents N7-methyl guanosine connected to the 5′ nucleotide through a 5′ to 5′ triphosphate linkage, and is also known as m7G cap or m7Gppp cap. The N7-MTase domain maps to the carboxy-terminal part of nsp14 (Chen et al. 2009). Cap-0 formation requires three sequential reactions catalyzed by RNA triphosphatase (TPase), guanylyltransferase (GTase), and N7-MTase. There is no evidence that nsp14 possesses TPase and GTase activities, and no other SARS-CoV-1 proteins with these activities have been identified, so the identities of the enzymes that mediate these required steps remain unknown. Based on the study of the human coronavirus 229E, non-structural protein 13 (nsp13) may have a TPase activity in addition to its established helicase activity (Ivanov and Ziebuhr 2004).
Nucleoprotein (N) is ADP-ribosylated. The modification is maintained both in the cell and in virions (Grunewald et al, 2018). Members of the protein mono-ADP-ribosyltransferase (PARP) enzyme family are thought to catalyze this reaction (Fehr et al. 2020)
Trimmed palmitoylated Spike protein trimers become associated with the ERGIC (ER-Golgi Intermediate Compartment) (Fung & Liu 2019)
A minor proportion of the E protein is modified by N-linked glycosylation. This variant appears to be more likely to form multimers, and it shows a different membrane topology than the main variant (Yuan et al, 2006).
In the presence of functional nsp14, which acts as a 3'-to-5' exonuclease, the mutation rate during human SARS coronavirus 1 (SARS-CoV-1) replication is 9 x 10^-7 (9E-7) per nucleotide per replication cycle or 2.2 x 10^-5 (2.2E-5) non-redundant substitutions per nucleotide, which translates into 2-3 nucleotide substitutions for each replicated SARS-CoV-1 genome. When nsp14 is defective, the mutation rate during SARS-CoV-1 replication increases to 1.2 x 10^-5 (1.2E-5) mutations per nucleotide per replication cycle or 3.34 x 10^-4 (3.34E-4) non-redundant substitutions per nucleotide, which translates into 12-23 nucleotide substitutions for each replicated SARS-CoV-1 genome (Eckerle et al. 2010). Here the process is annotated in two steps, nsp12- mediated misincorporation of a base (this reaction) and nsp14-mediated detection and removal of that base (next reaction).
The replicase polyprotein 1a of the human severe acute respiratory syndrome coronavirus (SARS-CoV) is post-translationally cleaved by virally encoded proteases to generate non-structural proteins (nsps). Viral nsps induce the formation of ER-bound double membrane vesicles (DMV) in host cells post infection. These DMVs are decorated with microtubule-associated proteins 1A/1B light chain 3B (MAP1LC3B) proteins that are involved in autophagosome formation. However, there is no evidence that DMVs are recruited to the autophagy machinery. Immunofluorescence studies show that nsp6 (Cottam E M. et al 2011) and nsp8 (Prentice E. et al 2004) colocalizes with MAP1LC3B suggesting a binding event. In some cell types, expression of sars9b (9b) triggers the formation of autophagosomes and underlying molecular mechanisms are unclear (Shi C S. et al 2014). Studies also show that sars8b (8b) can trigger cellular stress, which results in a calcineurin dependent Transcription Factor EB (TFEB) activation and its target genes. This leads to an increase in autophagic flux (Shi C S. et al 2019).
The uncoated virion nucleocapsid is released directly into the cytoplasm.The cellular protease furin cleaves the spike protein at the S1/S2 site and that cleavage is essential for S-protein-mediated cell-cell fusion and entry into human lung cells (Hoffman et al. 2020).
Transmembrane protease serine 2 (TMPRSS2), associated with the plasma membrane of the host cell, mediates the hydrolytic cleavage of SARS-CoV-2 Spike (S) protein component of the viral membrane-associate S3:M:E:encapsidated SARS coronavirus genomic RNA: 7a:O-glycosyl 3a tetramer complex associated with ACE2 (Matsuyama et al. 2010; Glowacka et al. 2011; Shulla et al. 2011).
the cellular protease furin cleaves the spike protein at the S1/S2 site and that cleavage is essential for S-protein-mediated cell-cell fusion and entry into human lung cells (Hoffman et al. 2020). Furin is associated with the plasma membrane of the host cell, mediates the hydrolytic cleavage of SARS-CoV-2 Spike (S) protein component of the viral membrane-associate S3:M:E:encapsidated SARS coronavirus genomic RNA: 7a:O-glycosyl 3a tetramer complex associated with ACE2.
7a:O-glycosyl 3a
tetramerSARS-CoV-2 genomic RNA: 7a:O-glycosyl 3a
tetramer:glycosylated-ACE2SARS-CoV-2 genomic RNA: 7a:O-glycosyl 3a
tetramer:glycosylated-ACE2SARS-CoV-2 genomic RNA: 7a:O-glycosyl 3a
tetramer:glycosylated-ACE2SARS-CoV-2 genomic RNA: 7a:O-glycosyl 3a
tetramer:glycosylated-ACE2SARS-CoV-2 genomic RNA: 7a:O-glycosyl 3a
tetramer:glycosylated-ACE2SARS-CoV-2 genomic RNA: 7a:O-glycosyl 3a
tetramer:glycosylated-ACE2SARS-CoV-2 genomic RNA: 7a:O-glycosyl 3a
tetramer:glycosylated-ACE2SARS-CoV-2 genomic RNA: 7a:O-glycosyl
3a tetramerSARS-CoV-2 genomic RNA: 7a:O-glycosyl
3a tetramerSARS-CoV-2 genomic RNA: 7a:O-glycosyl
3a tetramerSARS-CoV-2 genomic RNA: 7a:O-glycosyl
3a tetramerSARS-CoV-2 genomic RNA:7a:O-glycosyl
3a tetramerSARS-CoV-2 genomic RNA:7a:O-glycosyl
3a tetramerSARS-CoV-2 genomic RNA:O-glycosyl 3a
tetramerSARS-CoV-2 genomic RNA:O-glycosyl 3a
tetramergRNA with secondary
structure:RTCgRNA with secondary
structure:RTCgRNA with secondary
structure:RTCgRNA:RTC:nascent RNA minus strand with mismatched
nucleotidegRNA:RTC:nascent RNA minus strand with mismatched
nucleotidegRNA:RTC:nascent RNA minus strand with mismatched
nucleotidegRNA:RTC:RNA primer:RTC
inhibitorsgRNA:RTC:RNA primer:RTC
inhibitorsgRNA:RTC:RNA primer:RTC
inhibitorsgRNA:RTC:nascent RNA minus strand:RTC
inhibitorsgRNA:RTC:nascent RNA minus strand:RTC
inhibitorsgRNA:RTC:nascent
RNA minus strandgRNA:RTC:nascent
RNA minus strandgRNA:RTC:nascent
RNA minus strandgRNA:RTC:nascent
RNA minus strandgRNA:RTC:nascent
RNA minus strandgRNA:RTC:nascent
RNA minus strandgRNA:RTC:nascent
RNA minus strandcomplement (minus strand):RTC:RTC
inhibitorscomplement (minus strand):RTC:RTC
inhibitorscomplement (minus
strand):RTCcomplement (minus
strand):RTCcomplement (minus
strand):RTCcomplement (minus
strand):RTCRNA complement
(minus strand)RNA complement
(minus strand)strand subgenomic
mRNAsstrand subgenomic
mRNAsstrand subgenomic
mRNAsstrand subgenomic
mRNAsdimer:SARS-CoV-2
genomic RNAdimer:SARS-CoV-2
genomic RNAN-glycan-PALM-Spike
trimerN-glycan-PALM-Spike
trimerSARS-CoV-2 genomic
RNA (plus strand)SARS-CoV-2 genomic
RNA (plus strand)SARS-CoV-2 genomic
RNA (plus strand)SARS-CoV-2 genomic
RNASARS-CoV-2 genomic
RNAN-glycan folded
SpikeN-glycan folded
SpikeN-glycan unfolded
SpikeN-glycan unfolded
SpikeN-glycan-PALM-Spike
trimerN-glycan-PALM-Spike
trimerN-glycan-PALM-Spike
trimerN-glycan-PALM-Spike
trimerSARS-CoV-2 genomic
RNA (plus strand)SARS-CoV-2 genomic
RNA (plus strand)SARS-CoV-2 genomic RNA complement
(minus strand)SARS-CoV-2 genomic RNA complement
(minus strand)SARS-CoV-2 genomic RNA complement
(minus strand)polyadenylated SARS-CoV-2 subgenomic mRNAs
(plus strand)polyadenylated SARS-CoV-2 genomic
RNA (plus strand)polyadenylated SARS-CoV-2 genomic
RNA (plus strand)polyadenylated SARS-CoV-2 genomic
RNA (plus strand)polyadenylated SARS-CoV-2 genomic
RNA (plus strand)polyadenylated SARS-CoV-2 genomic
RNA (plus strand)polyadenylated SARS-CoV-2 genomic
RNA (plus strand)polyadenylated SARS-CoV-2 genomic
RNA (plus strand)polyadenylated SARS-CoV-2 genomic
RNA (plus strand)plus strand
subgenomic mRNAsSARS-CoV-2 genomic
RNA (plus strand)SARS-CoV-2 genomic
RNA (plus strand)SARS-CoV-2 genomic RNA complement
(minus strand)SARS-CoV-2 genomic RNA complement
(minus strand)N-glycan-PALM-Spike
trimerN-glycan-PALM-Spike
trimer