Determination of a Novel Epigenetic Silencing Mechanism



 DESCRIPTION (provided by applicant): The Epigenome regulates when, where and how an organism uses the genetic information stored in its genome. It is essential to many cellular processes, such as the regulation of gene expression, genome organization and cell-fate determination. It also governs growth, development, and ultimately human health. Heterochromatin represents silenced chromatic domains, which are assembled and maintained through epigenetic mechanisms. Extensive studies have focused on the assembly of heterochromatin at centromeres mediated by the RNA interference (RNAi) pathway. Recent studies, including ours, indicate that the exosome, an RNA quality control and surveillance complex, performs in a parallel pathway with RNAi to mediate epigenetic silencing. Although both RNAi and the exosome pathways are RNA-mediated and involved in processing long noncoding RNAs (ncRNAs) into small RNAs, exactly how the exosome pathway participates in heterochromatin assembly is not been demonstrated. Whether other RNAi-independent RNA processing pathways participate in epigenetic silencing is also unknown. We have recently identified a RNA-processing mechanism in epigenetic silencing that works independently of both RNAi and the exosome. The proposed research aims are designed to test the hypothesis that Dhp1, a conserved 5' to 3' exoribonuclease and ortholog of budding yeast Rat1 and metazoan Xrn2, plays a heretofore unknown role in epigenetic silencing, beyond its established role in transcription termination. Genetic, cell biology, and genomic approaches will be employed to investigate: 1) the localization of Dhp1 at heterochromatic regions; 2) Dhp1 binding proteins; and 3) the enzymatic activity of Dhp1 and potential co-activators in epigenetic silencing. Results will clarify how various RNA-processing pathways, acting together or independently, contribute to epigenetic regulation of the eukaryotic genome, a fundamental mechanism of gene expression. This study will be carried out in fission yeast, Schizosaccharomyces pombe (S. pombe), a premier model for studying epigenetic silencing because of its conserved epigenetic components and suitability for powerful combinatory experimental methods as well as undergraduate research. This project is significant and innovative. It will define a novel RNA-processing mechanism in epigenetic silencing that works independently of both RNAi and the exosome. Additionally, it will be the first exploration into how Dhp1/Rat1/Xrn2 functions in chromatin-based silencing.