The ability of a cell to arrest its growth is an essential function involved in numerous fundamental cellular processes. In contrast, unrestricted cell growth is a defining feature of cancer. Human oligoadenylate synthetase 1 (OAS1) is an interferon-inducible double-stranded RNA (dsRNA) binding protein that participates in several critical cellular processes requiring growth arrest, as well as in the innate immune response to viral infection. Whereas the activation of OAS1 by viral dsRNAs is well studied, the endogenous activation of OAS1 and its contribution to the regulation of cell growth remain largely uncharacterized. Interestingly, OAS1 is expressed as several alternatively spliced isoforms, which are present at varying levels in the population and which differentially correlate with disease. The proposed research will define the contributions of the OAS1 isoforms to physiological function and identify endogenous dsRNAs that activate OAS1 under a variety of growth-arrested cellular states in melanocytes and melanoma cells. Melanocytic cell lines present an ideal model for studying OAS1 function in normal and cancerous cells, because they can be induced to different cell states using physiologically relevant stimuli. First, rigorous methods will be developed to induce different growth-arrested cell states and quantitatively measure OAS1 activation. Next, the active isoform(s) in each growth-arrested state will be identified and their role in eliciting that particular cellular state will be determined using isofrm-specific depletion assays. Finally, deep sequencing will be used to identify endogenous dsRNAs that bind to immunoprecipitated OAS1 during growth-arrest, and the biochemical activities and physiological consequences of OAS1 binding to these dsRNAs will be characterized using both in vitro and cell culture- based assays. These studies will provide critical new insight into the relationship between OAS1 and essential cellular processes. Furthermore, this work will define novel roles for human dsRNAs in cell function and delineate how OAS1 participates in the aberrant growth characteristics of cancer.
Cells depend on an ability to stop growing in order to function properly, and while cancers may be associated with any number of genetic and pathological problems, cancer progression is always associated with unchecked growth. Activation of the double-stranded RNA-binding protein OAS1 results in the inhibition of cellular growth that is required during many critical cellular processes. This proposal will define the roles of different forms of human OAS1 and their RNA activators in normal cell function and in cancer cell pathology.
