UC San Diego

Nascent transcripts produced by RNA polymerase II in eukaryotic cells are subject to extensive processing prior to the generation of a functional messenger RNA (mRNA). These RNAs are generally found coated with RNA binding proteins (RBPs), which act in concert to regulate RNA processing events, including splicing, polyadenylation, and RNA stability. The aggregate effect of various RBPs on a given RNA transcript eventually dictates its fate, a phenomenon referred to as the RNP code. The precise control of RNA processing by RBPs is incredibly important for cellular homeostasis, defects of which lead to numerous accounts of human genetic diseases in many tissues. With the emergence of genome-wide methods for detecting direct binding of RBPs on target RNAs, coupled with technologies to measure changes in various aspects of RNA processing, global rules and insights for individual RBPs have been revealed. However, few studies have combined regulatory changes for more than one RBP to better understand their combinatorial effects on RNA targets. Additionally, despite the importance of RBPs, we still do not have the complete repertoire of which proteins bind to RNA, and which of these bind simultaneously with other RBPs to constitute the "RBP-RNA interactome". Here, I conduct genome-wide RNA processing analyses in mammalian cells, integrating regulation and binding information for multiple RBPs, including disease- related RBPs, the highly abundant heterogeneous nuclear ribonucleoparticle (hnRNP) proteins, and many previously uncharacterized RBPs. This battery of computational and experimental assays provides insight into the unique roles of hundreds of individual RBPs, as well as the extent of coordinated regulation between RBPs. I also describe a systematic approach to identifying the proteins that interact with RNA simultaneously with hnRNP proteins. Not limited to only mRNA-bound proteins, my strategy identifies thousands of hnRNP protein interactors, including putative novel proteins that interact with mRNA and pre-mRNA, and have biochemical and statistical attributes of known RBPs. My findings expand the repertoire of RNA-interacting RBPs and provide a resource for the study of human simultaneous RBP-RBP interactomes. This comprehensive analysis of RBPs investigates their specific roles in the regulation of RNA processing yielding interesting findings for the RNA biology field and insights into how misregulation can impact human disease