Decoding Repression: Recruitment of epigenetic silencers by RNA binding proteins and long non-coding RNAs - I aspire to be a principal investigator at a research-focused institution where I can study how RNA mediates gene regulation during early development. To this end, the activities proposed in this fellowship were designed to provide me with training in mechanistic aspects of RNA biology, developmental biology, computational analysis, scientific writing, and oral communication, which together will play essential roles in helping me establish a career in academic research. The overarching goal of my research is to delineate mechanisms by which long non- coding RNAs (lncRNAs) recruit chromatin-modifying (i.e., epigenetic) enzymes to regulate gene expression. Every step of development relies on dynamic gene regulation. As such, understanding how cells direct epigenetic enzymes to specific loci is essential to untangling the mechanisms that define early development. It has become clear that recruitment of epigenetic modifiers can be mediated by lncRNAs, the most potent of which, Xist, silences one of two X chromosomes in a process called X chromosome inactivation. However, it is not clear how lncRNAs encode the ability to recruit epigenetic modifiers. As the most powerfully repressive lncRNA known, Xist is an ideal model for decoding how lncRNAs recruit epigenetic modifiers and serves as a paradigm to understand other lncRNA-enzyme relationships. Xist-mediated silencing enzyme recruitment requires RBPs that are abundant in the cell, such as heterogeneous nuclear ribonucleoproteins (hnRNPs). Yet, paradoxically, hnRNPs bind thousands of other RNAs without contributing to transcriptional repression. The underlying sequence features and molecular interactions that allow Xist to exploit non-repressive RBPs to recruit epigenetic modifiers remain elusive. By focusing on a member of the hnRNP family, hnRNPK, which Xist requires to recruit the silencing enzyme complex Polycomb Repressive Complex 1 (PRC1), my research will 1) define the RNA sequence features that enable RNA to recruit PRC1 to chromatin and 2) determine how RNA and intrinsically disordered protein domains promote interactions between hnRNPK and PRC1. By defining the underlying RNA sequence features and the molecular interactions that enable PRC1 recruitment by RNA, I will identify a paradigm that will guide studies of other lncRNAs, RBPs, and silencing enzymes, which themselves are critical for embryonic development. These experiments will provide critical training in RNA biochemistry, genomics, computational biology, and quantitative microscopy, essential skills for studying RNA-mediated molecular mechanisms.
