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Principal Investigator: Jared Paul May
Organization: UNIVERSITY OF MISSOURI KANSAS CITY
Fiscal Year: 2023
Award: $78,250
Funding agency: National Institute of Allergy and Infectious Diseases
This proposal will investigate the role severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) N protein phase separation in disrupting cellular translation and RNA metabolism. N protein phase separation creates membraneless organelles and concentrates RNAs in a droplet (dense) phase. While N protein is responsible for encapsidation of the viral genome, N protein also binds hundreds of host messenger RNAs, some of which are necessary for translation and nonsense-mediated decay (NMD). NMD targets diverse families of RNA viruses for RNA decay, including betacoronaviruses, and the N protein from Murine hepatitis virus (MHV) interferes with NMD during the early stages of infection. Since NMD targets 10% of cellular transcripts for degradation, inhibition by N protein likely has profound effects on the host cell transcriptome. While the mechanism underlying the inhibition of NMD by N protein remains unclear, phase separation and translational repression by N could be a predominant factor. The cellular proteins FUS and C9orf72 arginine- rich dipeptide repeats are known to phase separate, repress translation, inhibit NMD, and are associated with Amyotrophic lateral sclerosis (ALS) disease progression. Viral proteins, including N protein, may repress the translation of transcripts necessary for the co-translational NMD pathway, potentially shaping the host transcriptome in a way that favors virus replication. Recent work has shown that SARS-CoV-2 infection induces high expression of inflammatory cytokines but fails to mount a robust interferon response. The underlying factors driving these transcriptional responses remain unknown; however, N protein could provoke this response by interfering with NMD and blocking post-transcriptional regulation. This project will use transcriptome-wide ribosome profiling (Ribo-seq) to identify cellular transcripts that are translationally repressed by N protein. Previously identified NMD targets will be examined to determine if translational repression confers NMD-resistance. Phase-separation deficient N mutants will be expressed in parallel to determine whether phase separation is the driving force in translation and NMD inhibition. Finally, the results generated from this study will be cross-referenced with published RNA-seq datasets from SARS-CoV-2 infected cells to determine the extent that N protein contributes to the large-scale changes in gene expression during infection. These findings will better our understanding of the virus-host interactions that take place during SARS-CoV-2 infection and will support on-going vaccine and antiviral development, especially N-based approaches.
Terms: <2019 novel corona virus><2019 novel coronavirus><2019-nCoV><Amyotrophic Lateral Sclerosis><Amyotrophic Lateral Sclerosis Motor Neuron Disease><Arginine><Automobile Driving><Basic Research><Basic Science><Binding><C-terminal><C9ORF72><COVID crisis><COVID epidemic><COVID infected patient><COVID pandemic><COVID patient><COVID positive patient><COVID-19><COVID-19 crisis><COVID-19 epidemic><COVID-19 global health crisis><COVID-19 global pandemic><COVID-19 health crisis><COVID-19 infected patient><COVID-19 infection><COVID-19 pandemic><COVID-19 patient><COVID-19 positive patient><COVID-19 public health crisis><COVID-19 virus><COVID-19 virus infection><COVID19><COVID19 crisis><COVID19 epidemic><COVID19 global health crisis><COVID19 global pandemic><COVID19 health crisis><COVID19 infection><COVID19 pandemic><COVID19 patient><COVID19 positive patient><COVID19 public health crisis><COVID19 virus><CV-19><CV19><Cell Body><Cells><Cessation of life><CoV-2><CoV2><Coronaviridae><Coronavirus><Data Set><Death><Development><Dipeptides><Disease><Disease Outbreaks><Disease Progression><Disorder><Event><FUS Protein><Family><Fusion Protein in Myxoid Liposarcoma><Future><G3BP><G3BP1><G3BP1 gene><Gehrig's Disease><Gene Expression><Gene Transcription><Genetic Transcription><Green Fluorescent Proteins><HeLa><Hela Cells><High-Throughput Nucleotide Sequencing><High-Throughput Sequencing><IFN><Immunoassay><Infection><Inflammatory><Interferons><L-Arginine><Length><Life Cycle><Life Cycle Stages><Lou Gehrig Disease><MERS corona virus><MERS coronavirus><MERS virus><MERS-CoV><Mediating><Membrane><Messenger RNA><Middle East Respiratory Syndrome Corona Virus><Middle East Respiratory Syndrome Coronavirus><Middle East Respiratory Syndrome Virus><Middle East Respiratory Syndrome-CoV><Middle East Respiratory coronavirus><Middle Eastern Respiratory Syndrome Corona virus><Middle Eastern Respiratory Syndrome Coronavirus><Middle Eastern Respiratory Syndrome Virus><Middle Eastern Respiratory Syndrome-CoV><Molecular Interaction><Mouse Hepatitis Virus><Murine Gastroenteritis Virus><Murine hepatitis virus><N Domain><Nature><Non-Polyadenylated RNA><Non-sense Mediated Decay><Nonsense-Mediated Decay><Nucleoproteins><Organelles><Outbreaks><POMp75 Protein><Pathway interactions><Phase><Phosphorylation><Post-Transcriptional Control><Post-Transcriptional Regulation><Process><Protein Phosphorylation><Proteins><Publishing><RNA><RNA Decay><RNA Degradation><RNA Expression><RNA Gene Products><RNA Seq><RNA Viruses><RNA metabolism><RNA sequencing><RNA-Binding Protein FUS><RNAseq><Reporter><Repression><Resistance><Ribo-seq><Ribonucleic Acid><Ribosomal RNA><Role><SARS Virus><SARS corona virus><SARS corona virus 2><SARS coronavirus><SARS-Associated Coronavirus><SARS-CO-V2><SARS-COVID-2><SARS-CoV><SARS-CoV-1><SARS-CoV-2><SARS-CoV-2 epidemic><SARS-CoV-2 global health crisis><SARS-CoV-2 global pandemic><SARS-CoV-2 infected patient><SARS-CoV-2 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patient><coronavirus disease-19 virus><coronavirus infectious disease-19><coronavirus patient><cytokine><design><designing><developing anti-viral agent><developing anti-viral drug><developing anti-viral therapeutic><developing anti-viral therapy><developing antiviral agent><developing antiviral drug><developing antiviral therapeutic><developing antiviral therapy><developmental><differential expression><differentially expressed><driving><driving force><effective therapy><effective treatment><global gene expression><global transcription profile><hCoV19><hnRNP P2><improved><infected with COVID-19><infected with COVID19><infected with SARS-CoV-2><infected with SARS-CoV2><infected with coronavirus disease 2019><infected with severe acute respiratory syndrome coronavirus 2><interfering virus><life course><mRNA><mRNA Stability><membrane structure><mutant><nCoV2><pathway><patient infected with COVID><patient infected with COVID-19><patient infected with SARS-CoV-2><patient infected with 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