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The EMBO Journal Oct 2020COVID-19, caused by SARS-CoV-2, has resulted in severe and unprecedented economic and social disruptions in the world. Nucleocapsid (N) protein, which is the major...
COVID-19, caused by SARS-CoV-2, has resulted in severe and unprecedented economic and social disruptions in the world. Nucleocapsid (N) protein, which is the major structural component of the virion and is involved in viral replication, assembly and immune regulation, plays key roles in the viral life cycle. Here, we solved the crystal structures of the N- and C-terminal domains (N-NTD and N-CTD) of SARS-CoV-2 N protein, at 1.8 and 1.5 Å resolution, respectively. Both structures show conserved features from other CoV N proteins. The binding sites targeted by small molecules against HCoV-OC43 and MERS-CoV, which inhibit viral infection by blocking the RNA-binding activity or normal oligomerization of N protein, are relatively conserved in our structure, indicating N protein is a promising drug target. In addition, certain areas of N-NTD and N-CTD display distinct charge distribution patterns in SARS-CoV-2, which may alter the RNA-binding modes. The specific antigenic characteristics are critical for developing specific immune-based rapid diagnostic tests. Our structural information can aid in the discovery and development of antiviral inhibitors against SARS-CoV-2 in the future.
Topics: Antiviral Agents; Betacoronavirus; Coronavirus Nucleocapsid Proteins; Crystallography, X-Ray; Drug Delivery Systems; Drug Design; Humans; Models, Molecular; Nucleocapsid Proteins; Phosphoproteins; Protein Conformation; Protein Domains; Recombinant Proteins; SARS-CoV-2
PubMed: 32914439
DOI: 10.15252/embj.2020105938 -
Cell Reports Jan 2023The leap of retroviruses and coronaviruses from animal hosts to humans has led to two ongoing pandemics and tens of millions of deaths worldwide. Retrovirus and... (Review)
Review
The leap of retroviruses and coronaviruses from animal hosts to humans has led to two ongoing pandemics and tens of millions of deaths worldwide. Retrovirus and coronavirus nucleocapsid proteins have been studied extensively as potential drug targets due to their central roles in virus replication, among which is their capacity to bind their respective genomic RNAs for packaging into nascent virions. This review focuses on fundamental studies of these nucleocapsid proteins and how their intrinsic abilities to condense through liquid-liquid phase separation (LLPS) contribute to viral replication. Therapeutic targeting of these condensates and methodological advances are also described to address future questions on how phase separation contributes to viral replication.
Topics: Humans; Coronavirus Nucleocapsid Proteins; COVID-19; Nucleocapsid Proteins; SARS-CoV-2; Virus Replication; HIV-1
PubMed: 36640305
DOI: 10.1016/j.celrep.2022.111968 -
Wiley Interdisciplinary Reviews. RNA 2016Viral nucleocapsid proteins (NCs) enwrap the RNA genomes of viruses to form NC-RNA complexes, which act as a template and are essential for viral replication and... (Review)
Review
Viral nucleocapsid proteins (NCs) enwrap the RNA genomes of viruses to form NC-RNA complexes, which act as a template and are essential for viral replication and transcription. Beyond packaging viral RNA, NCs also play important roles in virus replication, transcription, assembly, and budding by interacting with viral and host cellular proteins. Additionally, NCs can inhibit interferon signaling response and function in cell stress response, such as inducing apoptosis. Finally, NCs can be the target of vaccines, benefiting from their conserved gene sequences. Here, we summarize important findings regarding the additional functions of NCs as much more than structural RNA-binding proteins, with specific emphasis on (1) their association with the viral life cycle, (2) their association with host cells, and (3) as ideal candidates for vaccine development.
Topics: Animals; Gene Expression Regulation, Viral; Host-Pathogen Interactions; Humans; Immunity, Innate; Immunomodulation; Nucleocapsid Proteins; RNA Stability; RNA, Viral; Viral Vaccines; Virus Assembly; Virus Physiological Phenomena; Virus Replication
PubMed: 26749541
DOI: 10.1002/wrna.1326 -
Viruses Jul 2020Negative strand RNA viruses (NSVs) include many important human pathogens, such as influenza virus, Ebola virus, and rabies virus. One of the unique characteristics that... (Review)
Review
Negative strand RNA viruses (NSVs) include many important human pathogens, such as influenza virus, Ebola virus, and rabies virus. One of the unique characteristics that NSVs share is the assembly of the nucleocapsid and its role in viral RNA synthesis. In NSVs, the single strand RNA genome is encapsidated in the linear nucleocapsid throughout the viral replication cycle. Subunits of the nucleocapsid protein are parallelly aligned along the RNA genome that is sandwiched between two domains composed of conserved helix motifs. The viral RNA-dependent-RNA polymerase (vRdRp) must recognize the protein-RNA complex of the nucleocapsid and unveil the protected genomic RNA in order to initiate viral RNA synthesis. In addition, vRdRp must continuously translocate along the protein-RNA complex during elongation in viral RNA synthesis. This unique mechanism of viral RNA synthesis suggests that the nucleocapsid may play a regulatory role during NSV replication.
Topics: Genome, Viral; Models, Molecular; Negative-Sense RNA Viruses; Nucleocapsid; Nucleocapsid Proteins; Protein Conformation; Protein Folding; RNA, Viral; RNA-Dependent RNA Polymerase
PubMed: 32751700
DOI: 10.3390/v12080835 -
Vaccine Aug 2021The emergence and subsequent global outbreak of the novel coronavirus SARS-CoV-2 prompted our laboratory to launch efforts to develop methods for SARS-CoV-2 antigen...
The emergence and subsequent global outbreak of the novel coronavirus SARS-CoV-2 prompted our laboratory to launch efforts to develop methods for SARS-CoV-2 antigen detection and quantification. We present an isotope dilution mass spectrometry method (IDMS) for rapid and accurate quantification of the primary antigens, spike and nucleocapsid proteins. This IDMS method utilizes liquid chromatography-tandem mass spectrometry (LC-MS/MS) to analyze sample tryptic digests for detection and quantification of selected conserved peptides of SARS-CoV-2 spike and nucleocapsid proteins. The IDMS method has the necessary attributes to be successfully utilized for accurate quantification in SARS-CoV-2 protein-based vaccines and as targets of rapid diagnostic tests. Absolute quantification was achieved by quantifying and averaging 5 peptides for spike protein (3 peptides in the S1 subunit and 2 peptides in the S2 subunit) and 4 peptides for nucleocapsid protein. The overall relative standard deviation of the method was 3.67% for spike protein and 5.11% for nucleocapsid protein. IDMS offers speed (5 h total analysis time), sensitivity (LOQ; 10 fmol/µL) and precision for quantification of SARS-CoV-2 spike and nucleocapsid proteins.
Topics: COVID-19; Chromatography, Liquid; Coronavirus Nucleocapsid Proteins; Humans; Isotopes; Nucleocapsid Proteins; Phosphoproteins; SARS-CoV-2; Spike Glycoprotein, Coronavirus; Tandem Mass Spectrometry
PubMed: 34344552
DOI: 10.1016/j.vaccine.2021.07.066 -
Viruses Jul 2016The Bunyaviridae represents the largest family of segmented RNA viruses, which infect a staggering diversity of plants, animals, and insects. Within the family... (Review)
Review
The Bunyaviridae represents the largest family of segmented RNA viruses, which infect a staggering diversity of plants, animals, and insects. Within the family Bunyaviridae, the Phlebovirus genus includes several important human and animal pathogens, including Rift Valley fever virus (RVFV), severe fever with thrombocytopenia syndrome virus (SFTSV), Uukuniemi virus (UUKV), and the sandfly fever viruses. The phleboviruses have small tripartite RNA genomes that encode a repertoire of 5-7 proteins. These few proteins accomplish the daunting task of recognizing and specifically packaging a tri-segment complement of viral genomic RNA in the midst of an abundance of host components. The critical nucleation events that eventually lead to virion production begin early on in the host cytoplasm as the first strands of nascent viral RNA (vRNA) are synthesized. The interaction between the vRNA and the viral nucleocapsid (N) protein effectively protects and masks the RNA from the host, and also forms the ribonucleoprotein (RNP) architecture that mediates downstream interactions and drives virion formation. Although the mechanism by which all three genomic counterparts are selectively co-packaged is not completely understood, we are beginning to understand the hierarchy of interactions that begins with N-RNA packaging and culminates in RNP packaging into new virus particles. In this review we focus on recent progress that highlights the molecular basis of RNA genome packaging in the phleboviruses.
Topics: Animals; Humans; Insecta; Nucleocapsid Proteins; Phlebovirus; Plants; RNA, Viral; Virus Assembly
PubMed: 27428993
DOI: 10.3390/v8070194 -
Viruses Dec 2021Viruses of the family share a common and complex molecular machinery for transcribing and replicating their genomes. Their non-segmented, negative-strand RNA genome is... (Review)
Review
Viruses of the family share a common and complex molecular machinery for transcribing and replicating their genomes. Their non-segmented, negative-strand RNA genome is encased in a tight homopolymer of viral nucleoproteins (N). This ribonucleoprotein complex, termed a nucleocapsid, is the template of the viral polymerase complex made of the large protein (L) and its co-factor, the phosphoprotein (P). This review summarizes the current knowledge on several aspects of paramyxovirus transcription and replication, including structural and functional data on (1) the architecture of the nucleocapsid (structure of the nucleoprotein, interprotomer contacts, interaction with RNA, and organization of the disordered C-terminal tail of N), (2) the encapsidation of the genomic RNAs (structure of the nucleoprotein in complex with its chaperon P and kinetics of RNA encapsidation in vitro), and (3) the use of the nucleocapsid as a template for the polymerase complex (release of the encased RNA and interaction network allowing the progress of the polymerase complex). Finally, this review presents models of paramyxovirus transcription and replication.
Topics: Gene Expression Regulation, Viral; Humans; Nucleocapsid; Nucleocapsid Proteins; Paramyxoviridae Infections; Paramyxovirinae; Phylogeny; RNA, Viral
PubMed: 34960734
DOI: 10.3390/v13122465 -
Virology Nov 2019RNA viruses carry out selective packaging of their genomes in a variety of ways, many involving a genomic packaging signal. The first coronavirus packaging signal was... (Review)
Review
RNA viruses carry out selective packaging of their genomes in a variety of ways, many involving a genomic packaging signal. The first coronavirus packaging signal was discovered nearly thirty years ago, but how it functions remains incompletely understood. This review addresses the current state of knowledge of coronavirus genome packaging, which has mainly been studied in two prototype species, mouse hepatitis virus and transmissible gastroenteritis virus. Despite the progress that has been made in the mapping and characterization of some packaging signals, there is conflicting evidence as to whether the viral nucleocapsid protein or the membrane protein plays the primary role in packaging signal recognition. The different models for the mechanism of genomic RNA packaging that have been prompted by these competing views are described. Also discussed is the recent exciting discovery that selective coronavirus genome packaging is critical for in vivo evasion of the host innate immune response.
Topics: Models, Biological; Murine hepatitis virus; Nucleocapsid Proteins; RNA, Viral; Transmissible gastroenteritis virus; Viral Matrix Proteins; Virus Assembly
PubMed: 31505321
DOI: 10.1016/j.virol.2019.08.031 -
Science China. Life Sciences Apr 2012The nucleocapsid of vesicular stomatitis virus serves as the genomic template for transcription and replication. The viral genomic RNA is sequestered in the nucleocapsid... (Review)
Review
The nucleocapsid of vesicular stomatitis virus serves as the genomic template for transcription and replication. The viral genomic RNA is sequestered in the nucleocapsid in every step of the virus replication cycle. The structure of the nucleocapsid and the entire virion revealed how the viral genomic RNA is encapsidated and packaged in the virus. A unique mechanism for viral RNA synthesis is derived from the structure of the nuleocapsid and its interactions with the viral RNA-dependent RNA polymerase.
Topics: Microscopy, Electron; Models, Molecular; Nucleocapsid; Nucleocapsid Proteins; Protein Binding; RNA, Viral; RNA-Dependent RNA Polymerase; Vesicular stomatitis Indiana virus; Virus Assembly; Virus Replication
PubMed: 22566085
DOI: 10.1007/s11427-012-4307-x -
Antiviral Research Mar 2014The nucleocapsid phosphoprotein of the severe acute respiratory syndrome coronavirus (SARS-CoV N protein) packages the viral genome into a helical ribonucleocapsid (RNP)... (Review)
Review
The nucleocapsid phosphoprotein of the severe acute respiratory syndrome coronavirus (SARS-CoV N protein) packages the viral genome into a helical ribonucleocapsid (RNP) and plays a fundamental role during viral self-assembly. It is a protein with multifarious activities. In this article we will review our current understanding of the N protein structure and its interaction with nucleic acid. Highlights of the progresses include uncovering the modular organization, determining the structures of the structural domains, realizing the roles of protein disorder in protein-protein and protein-nucleic acid interactions, and visualizing the ribonucleoprotein (RNP) structure inside the virions. It was also demonstrated that N-protein binds to nucleic acid at multiple sites with a coupled-allostery manner. We propose a SARS-CoV RNP model that conforms to existing data and bears resemblance to the existing RNP structures of RNA viruses. The model highlights the critical role of modular organization and intrinsic disorder of the N protein in the formation and functions of the dynamic RNP capsid in RNA viruses. This paper forms part of a symposium in Antiviral Research on "From SARS to MERS: 10 years of research on highly pathogenic human coronaviruses."
Topics: Coronavirus Nucleocapsid Proteins; Humans; Macromolecular Substances; Models, Biological; Models, Molecular; Nucleocapsid Proteins; Phosphoproteins; Protein Binding; Protein Conformation; RNA, Viral; Severe acute respiratory syndrome-related coronavirus; Virus Assembly
PubMed: 24418573
DOI: 10.1016/j.antiviral.2013.12.009