Coronavirus nucleocapsid protein

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Nucleocapsid protein
SARS-CoV-2 (CDC-23312).png
Illustration of a SARS-CoV-2 virion.[1] The N protein, contained entirely within the virion, is not visible.
  Red: spike proteins (S)
  Yellow: envelope proteins (E)
  Orange: membrane proteins (M)
Identifiers
SymbolCoV_nucleocap
PfamPF00937
InterProIPR001218

The nucleocapsid (N) protein is a protein that packages the positive-sense RNA genome of coronaviruses to form ribonucleoprotein structures enclosed within the viral capsid.[2][3] The N protein is the most highly expressed of the four major coronavirus structural proteins.[2] In addition to its interactions with RNA, N forms protein-protein interactions with the coronavirus membrane protein (M) during the process of viral assembly.[2][3] N also has additional functions in manipulating the cell cycle of the host cell.[3][4] The N protein is highly immunogenic and antibodies to N are found in patients recovered from SARS and Covid-19.[5]

Structure[edit]

X-ray crystallography structure of the dimer formed by two C-terminal domains from the SARS-CoV-2 N protein.[6]

The N protein is composed of two main protein domains connected by an intrinsically disordered region (IDR) known as the linker region, with additional disordered segments at each terminus.[2][3] A third small domain at the C-terminal tail appears to have an ordered alpha helical secondary structure and may be involved in the formation of higher-order oligomeric assemblies.[6] In SARS-CoV, the causative agent of SARS, the N protein is 422 amino acid residues long[2] and in SARS-CoV-2, the causative agent of Covid-19, it is 419 residues long.[6][7]

Both the N-terminal and C-terminal domains are capable of binding RNA. The C-terminal domain forms a dimer that is likely to be the native functional state.[2] Parts of the IDR, particularly a conserved sequence motif rich in serine and arginine residues (the SR-rich region), may also be implicated in dimer formation, though reports on this vary.[2][3] Although higher-order oligomers formed through the C-terminal domain have been observed crystallographically, it is unclear if these structures have a physiological role.[2][8]

The C-terminal dimer has been structurally characterized by X-ray crystallography for several coronaviruses and has a highly conserved structure.[6] The N-terminal domain - sometimes known as the RNA-binding domain, though other parts of the protein also interact with RNA - has also been crystallized and has been studied by nuclear magnetic resonance spectroscopy in the presence of RNA.[9]

Post-translational modifications[edit]

The N protein is post-translationally modified by phosphorylation at sites located in the IDR, particularly in the SR-rich region.[2][10] In several coronaviruses, ADP-ribosylation of the N protein has also been reported.[11][10] With unclear functional significance, the SARS-CoV N protein has been observed to be SUMOylated and the N proteins of several coronaviruses have been observed to be proteolytically cleaved.[10]

Expression and localization[edit]

The N protein is the most highly expressed in host cells of the four major structural proteins.[2] Like the other structural proteins, the gene encoding the N protein is located toward the 3' end of the genome.[3]

N protein is localized primarily to the cytoplasm.[3] In many coronaviruses, a population of N protein is localized to the nucleolus,[3][4][12] thought to be associated with its effects on the cell cycle.[4]

Function[edit]

Genome packaging and viral assembly[edit]

Coronavirus virion structure cross-section. The N proteins are represented by the small circles associated with the RNA genome in the virion interior.
NMR structure of the SARS-CoV-2 N protein N-terminal domain (red) in complex with double-stranded RNA (orange and yellow).[9]

The N protein binds to RNA to form ribonucleoprotein (RNP) structures for packaging the genome into the viral capsid.[2][3] The RNP particles formed are roughly spherical and are organized in flexible helical structures inside the virus.[2][3] Formation of RNPs is thought to involve allosteric interactions between RNA and multiple RNA-binding regions of the protein.[2][8] Dimerization of N is important for assembly of RNPs. Encapsidation of the genome occurs through interactions between N and M.[2][3] N is essential for viral assembly.[3] N also serves as a chaperone protein for the formation of RNA structure in the genomic RNA.[3][8]

Genomic and subgenomic RNA synthesis[edit]

Synthesis of genomic RNA appears to involve participation by the N protein. N is physically colocalized with the viral RNA-dependent RNA polymerase early in the replication cycle and forms interactions with non-structural protein 3, a component of the replicase-transcriptase complex.[3] Although N appears to facilitate efficient replication of genomic RNA, it is not required for RNA transcription in all coronaviruses.[3][13] In at least one coronavirus, transmissible gastroenteritis virus (TGEV), N is involved in template switching in the production of subgenomic mRNAs, a process that is a distinctive feature of viruses in the order Nidovirales.[3][13][14]

Cell cycle effects[edit]

Coronaviruses manipulate the cell cycle of the host cell through various mechanisms. In several coronaviruses, including SARS-CoV, the N protein has been reported to cause cell cycle arrest in S phase through interactions with cyclin-CDK.[3][4] In SARS-CoV, a cyclin box-binding region in the N protein can serve as a cyclin-CDK phosphorylation substrate.[3] Trafficking of N to the nucleolus may also play a role in cell cycle effects.[4] More broadly, N may be involved in reduction of host cell protein translation activity.[3]

Immune system effects[edit]

The N protein is involved in viral pathogenesis via its effects on components of the immune system. In SARS-CoV,[3][15][16] MERS-CoV,[17] and SARS-CoV-2,[18] N has been reported as suppressing interferon responses.

Evolution and conservation[edit]

The structures of N proteins from different coronaviruses, particularly the C-terminal domains, appear to be well conserved.[2][6] Similarities between the structure and topology of the N proteins of coronaviruses and arteriviruses suggest a common evolutionary origin and supports the classification of these two groups in the common order Nidovirales.[2][3]

Examination of SARS-CoV-2 sequences collected during the Covid-19 pandemic found that missense mutations were most common in the central linker region of the protein, suggesting this relatively unstructured region is more tolerant of mutations than the structured domains.[6] A separate study of SARS-CoV-2 sequences identified at least one site in the N protein under positive selection.[19]

References[edit]

  1. ^ Giaimo C (1 April 2020). "The Spiky Blob Seen Around the World". The New York Times. Archived from the original on 2 April 2020. Retrieved 6 April 2020.
  2. ^ a b c d e f g h i j k l m n o p Chang, Chung-ke; Hou, Ming-Hon; Chang, Chi-Fon; Hsiao, Chwan-Deng; Huang, Tai-huang (March 2014). "The SARS coronavirus nucleocapsid protein – Forms and functions". Antiviral Research. 103: 39–50. doi:10.1016/j.antiviral.2013.12.009. PMC 7113676.
  3. ^ a b c d e f g h i j k l m n o p q r s t u McBride, Ruth; van Zyl, Marjorie; Fielding, Burtram (7 August 2014). "The Coronavirus Nucleocapsid Is a Multifunctional Protein". Viruses. 6 (8): 2991–3018. doi:10.3390/v6082991. PMC 4147684.
  4. ^ a b c d e Su, Mingjun; Chen, Yaping; Qi, Shanshan; Shi, Da; Feng, Li; Sun, Dongbo (5 November 2020). "A Mini-Review on Cell Cycle Regulation of Coronavirus Infection". Frontiers in Veterinary Science. 7: 586826. doi:10.3389/fvets.2020.586826. PMC 7674852.
  5. ^ Li, Dandan; Li, Jinming (20 April 2021). "Immunologic Testing for SARS-CoV-2 Infection from the Antigen Perspective". Journal of Clinical Microbiology. 59 (5). doi:10.1128/JCM.02160-20. PMC 8091849.
  6. ^ a b c d e f Ye, Qiaozhen; West, Alan M. V.; Silletti, Steve; Corbett, Kevin D. (September 2020). "Architecture and self‐assembly of the SARS‐CoV ‐2 nucleocapsid protein". Protein Science. 29 (9): 1890–1901. doi:10.1002/pro.3909.
  7. ^ Shah, Vibhuti Kumar; Firmal, Priyanka; Alam, Aftab; Ganguly, Dipyaman; Chattopadhyay, Samit (7 August 2020). "Overview of Immune Response During SARS-CoV-2 Infection: Lessons From the Past". Frontiers in Immunology. 11: 1949. doi:10.3389/fimmu.2020.01949. PMC 7426442.
  8. ^ a b c Chang, Chung-ke; Lo, Shou-Chen; Wang, Yong-Sheng; Hou, Ming-Hon (April 2016). "Recent insights into the development of therapeutics against coronavirus diseases by targeting N protein". Drug Discovery Today. 21 (4): 562–572. doi:10.1016/j.drudis.2015.11.015. PMC 7108309.
  9. ^ a b Dinesh, Dhurvas Chandrasekaran; Chalupska, Dominika; Silhan, Jan; Koutna, Eliska; Nencka, Radim; Veverka, Vaclav; Boura, Evzen (2 December 2020). "Structural basis of RNA recognition by the SARS-CoV-2 nucleocapsid phosphoprotein". PLOS Pathogens. 16 (12): e1009100. doi:10.1371/journal.ppat.1009100. PMC 7735635.
  10. ^ a b c Fung, To Sing; Liu, Ding Xiang (June 2018). "Post-translational modifications of coronavirus proteins: roles and function". Future Virology. 13 (6): 405–430. doi:10.2217/fvl-2018-0008. PMC 7080180.
  11. ^ Grunewald, Matthew E.; Fehr, Anthony R.; Athmer, Jeremiah; Perlman, Stanley (April 2018). "The coronavirus nucleocapsid protein is ADP-ribosylated". Virology. 517: 62–68. doi:10.1016/j.virol.2017.11.020. PMC 5871557.
  12. ^ Masters, Paul S. (2006). "The Molecular Biology of Coronaviruses". Advances in Virus Research. 66: 193–292. doi:10.1016/S0065-3527(06)66005-3. PMC 7112330.
  13. ^ a b Zúñiga, Sonia; Cruz, Jazmina L. G.; Sola, Isabel; Mateos-Gómez, Pedro A.; Palacio, Lorena; Enjuanes, Luis (15 February 2010). "Coronavirus Nucleocapsid Protein Facilitates Template Switching and Is Required for Efficient Transcription". Journal of Virology. 84 (4): 2169–2175. doi:10.1128/JVI.02011-09. PMC 2812394.
  14. ^ Sola, Isabel; Almazán, Fernando; Zúñiga, Sonia; Enjuanes, Luis (9 November 2015). "Continuous and Discontinuous RNA Synthesis in Coronaviruses". Annual Review of Virology. 2 (1): 265–288. doi:10.1146/annurev-virology-100114-055218. PMC 6025776.
  15. ^ Spiegel, Martin; Pichlmair, Andreas; Martínez-Sobrido, Luis; Cros, Jerome; García-Sastre, Adolfo; Haller, Otto; Weber, Friedemann (15 February 2005). "Inhibition of Beta Interferon Induction by Severe Acute Respiratory Syndrome Coronavirus Suggests a Two-Step Model for Activation of Interferon Regulatory Factor 3". Journal of Virology. 79 (4): 2079–2086. doi:10.1128/JVI.79.4.2079-2086.2005. PMC 546554.
  16. ^ Kopecky-Bromberg, Sarah A.; Martínez-Sobrido, Luis; Frieman, Matthew; Baric, Ralph A.; Palese, Peter (15 January 2007). "Severe Acute Respiratory Syndrome Coronavirus Open Reading Frame (ORF) 3b, ORF 6, and Nucleocapsid Proteins Function as Interferon Antagonists". Journal of Virology. 81 (2): 548–557. doi:10.1128/JVI.01782-06. PMC 1797484.
  17. ^ Chang, Chi-You; Liu, Helene Minyi; Chang, Ming-Fu; Chang, Shin C. (16 June 2020). "Middle East Respiratory Syndrome Coronavirus Nucleocapsid Protein Suppresses Type I and Type III Interferon Induction by Targeting RIG-I Signaling". Journal of Virology. 94 (13). doi:10.1128/JVI.00099-20. PMC 7307178.
  18. ^ Mu, Jingfang; Fang, Yaohui; Yang, Qi; Shu, Ting; Wang, An; Huang, Muhan; Jin, Liang; Deng, Fei; Qiu, Yang; Zhou, Xi (December 2020). "SARS-CoV-2 N protein antagonizes type I interferon signaling by suppressing phosphorylation and nuclear translocation of STAT1 and STAT2". Cell Discovery. 6 (1): 65. doi:10.1038/s41421-020-00208-3. PMC 7490572.
  19. ^ Cagliani, Rachele; Forni, Diego; Clerici, Mario; Sironi, Manuela (June 2020). "Computational Inference of Selection Underlying the Evolution of the Novel Coronavirus, Severe Acute Respiratory Syndrome Coronavirus 2". Journal of Virology. 94 (12). doi:10.1128/JVI.00411-20. PMC 7307108.