Document text
Principal Investigator: Beatriz MA Fontoura
Organization: VANDERBILT UNIVERSITY
Fiscal Year: 2024
Award: $857,199
Funding agency: National Institute of Allergy and Infectious Diseases
The global COVID-19 pandemic caused by the SARS-CoV-2 outbreak underscores the need for understanding viral-host interactions that impact host gene expression and antiviral responses. A critical virulence factor of coronaviruses is the nonstructural protein 1 (Nsp1). We uncovered a previously unknown mechanism of Nsp1- mediated inhibition of host antiviral responses: Nsp1 targets the cellular mRNA nuclear export pathway. Nuclear export of cellular mRNAs through the nuclear pore complex (NPC) is obligatory for eukaryotic gene expression including those genes encoding antiviral factors. By contrast, SARS-CoV-2 mRNA synthesis takes place at viral transcriptional factories in the cytoplasm. We have demonstrated that SARS-CoV-2 Nsp1 protein inhibits host mRNA nuclear export by targeting the cellular NXF1-NXT1 complex, a receptor required for mRNA translocation through the NPC to the cytoplasm. We identified mutations in Nsp1 that specifically reduce its binding to NXF1- NXT1. Importantly, a SARS-CoV-2 recombinant virus carrying a mutant Nsp1 deficient in NXF1-NXT1 binding is unable to inhibit mRNA nuclear export and is attenuated in cells and in mice. Together, our data indicate that Nsp1 is a regulator of host mRNA nuclear export via blocking NXF1 function, overcoming cellular antiviral responses. This proposal combines the synergistic expertise between the principal investigators in biochemistry, cell biology, structural biology, and viral pathogenesis to investigate in depth the molecular mechanisms of SARS-CoV-2 Nsp1 protein function as inhibitor of mRNA nuclear export and its impact on the virus life cycle and on virulence. In Aim 1, we will use biochemical, genetic, and imaging approaches to determine the interactions within the mRNA export machinery that lead to Nsp1 inhibition of mRNA export. In addition, we will use a combination of biophysical and structural biology approaches to determine the molecular basis for the recognition between Nsp1 and NXF1-NXT1. These findings will uncover new mechanisms that mediate or regulate cellular mRNA nuclear export and how they are targeted by SARS-CoV-2. In Aim 2, we will assess the functions of the Nsp1-NXF1 interaction during virus replication and pathogenesis. We will generate loss-of-function recombinant viruses carrying Nsp1 mutants defective in NXF1 binding and investigate the impact of these mutations on host mRNA export, viral replication, induction of host responses in infected cells, and on viral pathogenesis in two animal models, mice and hamsters. Additionally, Nsp1 polymorphisms associated with new variants of concern have been found. We will therefore address the functional significance of these polymorphisms for the Nsp1- NXF1 interaction and for viral pathogenesis. Moreover, we will identify the host mRNAs whose nuclear export are impacted by Nsp1 and determine their role in antiviral response. These extensive studies will likely not only define novel virulence function(s) of SARS-CoV-2 Nsp1, but also shed light on novel antiviral defense mechanisms. Since antagonizing Nsp1 function results in expression of mRNAs encoding antiviral factors, our proposed studies may uncover new strategies for designing antivirals against SARS-CoV-2.
Terms: <2019 novel corona virus><2019 novel coronavirus><2019-nCoV><Address><Animal Model><Animal Models and Related Studies><Anti-viral Agents><Anti-viral Response><Assay><Attenuated><Binding><Binding Sites><Bioassay><Biochemical><Biochemistry><Biological><Biological Assay><Biological Chemistry><Biophysics><COVID crisis><COVID epidemic><COVID pandemic><COVID-19 crisis><COVID-19 epidemic><COVID-19 era><COVID-19 global health crisis><COVID-19 global pandemic><COVID-19 health crisis><COVID-19 infection><COVID-19 outbreak><COVID-19 pandemic><COVID-19 period><COVID-19 public health crisis><COVID-19 virus><COVID-19 virus infection><COVID-19 years><COVID19 infection><COVID19 outbreak><COVID19 virus><Cell Body><Cell Function><Cell Nucleus><Cell Physiology><Cell Process><Cells><Cellular Function><Cellular Physiology><Cellular Process><Cellular biology><CoV-2><CoV2><Combining Site><Complex><Coronaviridae><Coronavirus><Cricetinae><Cytoplasm><Cytoplasmic Protein><Data><Defense Mechanisms><Development><Gene Expression><Gene Transcription><Genes><Genetic Alteration><Genetic Change><Genetic Polymorphism><Genetic Transcription><Genetic defect><Hamsters><Hamsters Mammals><Immune response><Immunological response><Innate Immune Response><Knowledge><Laboratories><Life Cycle><Life Cycle Stages><Mediating><Messenger RNA><Mice><Mice Mammals><Molecular><Molecular Interaction><Murine><Mus><Mutation><NPC><Non-structural Protein><Nonstructural Protein><Nuclear><Nuclear Export><Nuclear Import><Nuclear Pore Complex><Nucleus><Pathogenesis><Pathogenicity><Pathogenicity Factors><Pathway interactions><Principal Investigator><Proteins><Proteomics><RNA Expression><Reactive Site><Receptor Protein><Recombinants><Regulation><Role><SARS corona virus 2><SARS-CO-V2><SARS-COVID-2><SARS-CoV-2><SARS-CoV-2 epidemic><SARS-CoV-2 global health crisis><SARS-CoV-2 global pandemic><SARS-CoV-2 infection><SARS-CoV-2 outbreak><SARS-CoV-2 pandemic><SARS-CoV-2 pathogenesis><SARS-CoV2><SARS-CoV2 infection><SARS-associated corona virus 2><SARS-associated coronavirus 2><SARS-coronavirus-2><SARS-coronavirus-2 epidemic><SARS-coronavirus-2 pandemic><SARS-related corona virus 2><SARS-related coronavirus 2><SARSCoV2><Severe Acute Respiratory Coronavirus 2><Severe Acute Respiratory Distress Syndrome CoV 2><Severe Acute Respiratory Distress Syndrome Corona Virus 2><Severe Acute Respiratory Distress Syndrome Coronavirus 2><Severe Acute Respiratory Syndrome CoV 2><Severe Acute Respiratory Syndrome CoV 2 epidemic><Severe Acute Respiratory Syndrome CoV 2 pandemic><Severe Acute Respiratory Syndrome-associated coronavirus 2><Severe Acute Respiratory Syndrome-related coronavirus 2><Severe acute respiratory syndrome associated corona virus 2><Severe acute respiratory syndrome coronavirus 2><Severe acute respiratory syndrome coronavirus 2 epidemic><Severe acute respiratory syndrome coronavirus 2 infection><Severe acute respiratory syndrome coronavirus 2 outbreak><Severe acute respiratory syndrome coronavirus 2 pandemic><Severe acute respiratory syndrome related corona virus 2><Site><Structure><Subcellular Process><Topical Drug Administration><Topical application><Transcription><Translations><Travel><Viral><Viral Gene Products><Viral Gene Proteins><Viral Pathogenesis><Viral Proteins><Virulence><Virulence Factors><Virus><Virus Replication><Wuhan coronavirus><administer topically><anti-viral compound><anti-viral drugs><anti-viral medication><anti-viral therapeutic><anti-virals><apply topically><attenuate><attenuates><biologic><biophysical foundation><biophysical principles><biophysical sciences><cell biology><cellular targeting><corona virus><coronavirus disease 2019 crisis><coronavirus disease 2019 epidemic><coronavirus disease 2019 global health crisis><coronavirus disease 2019 global pandemic><coronavirus disease 2019 health crisis><coronavirus disease 2019 infection><coronavirus disease 2019 outbreak><coronavirus disease 2019 pandemic><coronavirus disease 2019 public health crisis><coronavirus disease 2019 virus><coronavirus disease crisis><coronavirus disease epidemic><coronavirus disease pandemic><coronavirus disease-19 global pandemic><coronavirus disease-19 outbreak><coronavirus disease-19 pandemic><coronavirus disease-19 virus><deliver topically><design><designing><developmental><genetic approach><genetic strategy><genome mutation><hCoV19><host response><imaging approach><imaging based approach><immune system response><immunoresponse><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><inhibitor><insight><interdisciplinary approach><life course><loss of function><mRNA><mRNA Export><mRNA Expression><member><model of animal><multidisciplinary approach><mutant><nCoV2><new therapeutic approach><new therapeutic intervention><new therapeutic strategies><new therapy approaches><new treatment approach><new treatment strategy><novel><novel therapeutic approach><novel therapeutic intervention><novel therapeutic strategies><novel therapy approach><outbreak of SARS-CoV-2><pathway><polymorphism><protein function><psychological defense mechanism><receptor><recombinant virus><response><severe acute respiratory syndrome coronavirus 2 global health crisis><severe acute respiratory syndrome coronavirus 2 global pandemic><severe acute respiratory syndrome coronavirus 2 pathogenesis><social role><structural biology><topical administration><topical delivery><topical drug application><topical treatment><topically administered><topically applied><topically delivered><topically treated><translation><treat topically><variants of concern><viral multiplication><viral replication><virology><virus multiplication><virus pathogenesis><virus protein>