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Principal Investigator: Alison Whitney Ashbrook
Organization: ROCKEFELLER UNIVERSITY
Fiscal Year: 2020
Award: $69,306
Funding agency: National Institute of Allergy and Infectious Diseases
PROJECT SUMMARY/ABSTRACT
Translation is a critical stage at which the abundance, timing, and localization of gene expression is
regulated. In the case of infection, precise control of translational machinery and processes can determine
whether an effective antiviral program is initiated or whether these programs are impeded by the invading
pathogen. Although required for both virus replication and host defense, mechanisms by which translation is
regulated early after virus infection are not fully understood. Moreover, the importance of translational
regulation in limiting viral replication remains to be elucidated. Understanding functions for translational
regulation during virus infection and innate immune responses will provide insights into virus-induced disease
and uncover novel mechanisms of antiviral immunity. The main goal of this proposal is to investigate
functions of early regulation of translation in host susceptibility to virus infection. In Specific Aim 1, I
will use ribosome profiling and pulse-labeled mass spectrometry to identify and quantify translationally
regulated mRNAs in immune stimulated cells. Cells deficient for specific translationally regulated mRNAs will
be used to determine the importance of these mRNAs in viral replication and antiviral immunity. In Specific
Aim 2, I will utilize confocal microscopy of pulse-labeled cells to define subcellular sites of early translational
events that follow virus infection and immune activation. Translation of specific antiviral mRNAs also will be
examined by selective tagging and visualized by live-cell microscopy. In Specific Aim 3, I will study ribosomal
stalk proteins, which were recently identified as pro-viral host factors in a CRISPR knockout screen, and their
roles during virus infection. Expression and localization of the stalk proteins will be determined following
immune stimulation, and translation and viral replication will be assessed in stalk protein-deficient cells. Stalk
protein-deficient cells will be reconstituted with genetically altered constructs to define sequences that
contribute to stalk protein function. Together, these studies will elucidate functions for translational regulation in
host susceptibility to virus infection and will enhance our understanding of host-pathogen interactions.
Knowledge gained from this proposed research may illuminate new approaches for therapeutically targeting
broad families of viruses.
Terms: <Active Sites><Affect><Amino Acid Sequence><Antibodies><Antiviral Agents><Antiviral Drugs><Antivirals><Body Tissues><CRISPR><CRISPR/Cas system><Cell Body><Cell Communication and Signaling><Cell Signaling><Cells><Chromosome Mapping><Clustered Regularly Interspaced Short Palindromic Repeats><Complex><Confocal Microscopy><Detection><Development><Elongation Factor><Eukaryotic Cell><Event><Family><Flavivirus><Flavivirus Infections><Gene Expression><Gene Localization><Gene Mapping><Gene Mapping Genetics><Gene Transcription><Genes><Genetic Transcription><Genetic Translation><Goals><Group B Arbovirus><Host Defense><Host Defense Mechanism><Host Factor><Host Factor Protein><Human><IFN><Immune><Immune Cell Activation><Immune response><Immunes><Immunization><Immunologic Sensitization><Immunologic Stimulation><Immunological Sensitization><Immunological Stimulation><Immunological response><Immunostimulation><Individual><Infection><Innate Immune Response><Innate Immunity><Integration Host Factors><Interferons><Intracellular Communication and Signaling><Invaded><Kinetics><Knock-out><Knockout><Knowledge><Label><Linkage Mapping><Mass Photometry/Spectrum Analysis><Mass Spectrometry><Mass Spectroscopy><Mass Spectrum><Mass Spectrum Analyses><Mass Spectrum Analysis><Measures><Mediating><Messenger RNA><Modern Man><Monitor><Native Immunity><Natural Immunity><Nerve Cells><Nerve Unit><Neural Cell><Neurocyte><Neurons><Non-Polyadenylated RNA><Non-Specific Immunity><Nonspecific Immunity><Organelles><Pattern><Physiologic pulse><Predisposition><Primary Protein Structure><Process><Proteins><Pulse><RNA><RNA Expression><RNA Gene Products><Regulation><Research><Ribo-seq><Ribonucleic Acid><Ribosomal Proteins><Ribosomes><Role><Shapes><Signal Transduction><Signal Transduction Systems><Signaling><Site><Specificity><Susceptibility><Testing><Therapeutic><Tissues><Total Human and Non-Human Gene Mapping><Transcript><Transcription><Translating><Translational Regulation><Translations><Up-Regulation><Upregulation><Vaccine Design><Viral><Viral Diseases><Viral Pathogenesis><Virus><Virus Diseases><Virus Replication><Work><anti-viral agents><anti-viral drugs><anti-viral immunity><anti-virals><antiviral immunity><biological signal transduction><defined contribution><developmental><experiment><experimental research><experimental study><genetic mapping><host response><immune activation><immunoresponse><insight><live cell microscopy><mRNA><mRNA Translation><neuronal><new approaches><new therapeutic approach><new therapeutic intervention><new therapeutic strategies><new therapy approaches><novel><novel approaches><novel strategies><novel strategy><novel therapeutic approach><novel therapeutic intervention><novel therapy approach><pathogen><programs><protein function><protein sequence><reconstitute><reconstitution><recruit><response><ribosome footprint profiling><ribosome profiling><social role><spatiotemporal><therapeutic target><translatome><viral detection><viral infection><viral multiplication><viral replication><virus detection><virus infection><virus multiplication><virus pathogenesis><virus-induced disease>