Emerging pathogens, such as SARS-CoV, Emerging Coronavirus 2012 {EMC/2012), influenza virus (lAV), Filoviruses (Ebola) and Human Herpesvirus 8 {HHV8) are significant public health threats. All of these pathogens encode uncharacterized open reading frames (ORFs) and noncoding RNAs that we hypothesize will act as either positive or negative regulators of viral replication or modulators of host cell regulatory processes or antiviral responses. Therefore, investigating how these ORFs/RNAs affect viral replication and/or the host response will enhance our general understanding of the pathogenesis of each of these viruses and may identify common pathways that can be targeted for therapeutic intervention. Our program involves a highly interactive group of investigators, who will work collaboratively to characterize the impact of novel viral ORFs and/or noncoding RNAs in regulating viral replication or key host processes that are likely to be targets of viral modulation, such as type I IFN response, apoptosis, or inflammasome activation. This analysis requires that each research project has access to a standardized set of expression systems that allow for ready cross comparison of protein/RNA function between projects. Therefore, the core will clone and express candidate ORFs/noncoding RNAs from standard sets of expression vectors, perform quality control assays to ensure appropriate expression of the candidate ORF/RNA, and then provide these expression vectors to each of the research projects. Since one of the goals of this program is to characterize how novel ORFs affect viral pathogenesis, we will generate antisera against each of the candidate ORFs, validate the specificity of this antiserum, and then provide it to each project for use in assessing protein expression, cellular localization, and protein/protein interactions in the context of authentic SARS-CoV, EMC/2012, lAV, Ebola virus, or HHV 8 infection. Lastly, for each candidate ORF, the core will use the expression vectors to perform detailed analysis of the expression and biochemical characteristics of each ORF (e.g. posttranslational modifications such as glycosylation) in collaboration with each research project.
Core will interact closely with each research project to provide a set of standardized expression vectors encoding uncharacterized ORFs or noncoding RNAs. The core will also provide ORF specific antiserum and will biochemically characterize each of the uncharacterized ORFs in collaboration with the three research projects.
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