Post Transcriptional Gene Regulation By The Double Stranded Rna Binding Protein Staufen1 PDF Download

RNA binding proteins (RBPs) are key regulators of gene expression that, by interacting with specific RNAs, can impact their splicing, localization, translation and decay. Staufen proteins are RBPs that are conserved through metazoan evolution, and can simultaneously bind both double-stranded RNAs (dsRNAs) and diverse proteins. These properties, together with its presence in the nucleus, cytoplasm and at the endoplasmic reticulum (ER), has made it a versatile regulatory protein. In this thesis I explore three distinct contexts wherein Staufen1 has been co-opted during mammalian evolution to regulate gene expression. The first example pursues the Maquat-lab finding that mammalian Staufen1 (STAU1) binds dsRNA structures formed between primate-specific Alu short interspersed elements (SINEs) in humans and between rodent-specific B or ID (B/ID) SINEs in mice. When these duplexes are present in mRNA 3'-untranslated regions (3'-UTRs), STAU1 binding promotes Staufen-mediated decay (SMD). We hypothesized that gene regulation resulting from SINE insertion might have been similarly adaptive in separate mammalian lineages, leading to parallel evolution of the Staufen network by independent exaptation of SINEs. To explore this hypothesis, we identified and confirmed orthologous gene pairs with 3'-UTR SINEs that have been independently exapted in mouse and humans for SMD control of myoblast metabolism. Expanding to other species we demonstrated that SINE-directed SMD likely emerged in both primate and rodent lineages>25 million years ago. This work reveals a novel mechanism for the convergent evolution of post-transcriptional gene regulatory networks in mammals by species-specific SINE transposition and SMD. Another consequence of STAU1 binding to duplexes formed between inverted Alu repeats (IRAlus) in mRNA 3'-UTRs is to promote the escape from nuclear retention. While some mRNAs harboring 3'-UTR IRAlus can be retained in the nucleus by interaction with the paraspeckle protein p54nrb, other 3'-UTR IRAlus do not promote nuclear retention but do promote mRNA translation. The diverse consequences of 3'-UTR Alus imply the existence of different, as yet undefined subcategories. To reach a global understanding of the contribution of 3'-UTR Alus to mRNA localization, translation and decay we undertook to characterize the nucleocytoplasmic distribution, translation and abundance of the 3'-UTR Alu element-containing transcriptome using the isoform-specific RNA seq method, READS+. Using this method uncovered 3'-UTR features that correlate with mRNA localization and distinguish a subset of IRAlus-containing mRNAs, for which the mRNA export is regulated by STAU1. We made an additional and unexpected finding that 3'-UTR length is a determinant of mRNA localization, independent of Alu content. STAU1 also binds intramolecular duplexes in ER-localized mRNAs, including the ER stress-responsive transcript XBP1. Under ER stress, this transcript is known to undergo cytoplasmic cleavage and ligation resulting in a translational frameshift and production of a potent transcription factor that activates ER stress-responsive genes. I have show that knockdown of Staufen1 reduced the production of spliced XBP1 in response to ER stress without a concomitant increase in unspliced XBP1, which, together with my finding that STAU1 co-immunoprecipitates with the RNA ligase RTCB, implicates Staufen1 as a regulator of the ligation step, rather than the cleavage step, of XBP1 splicing. Depletion of Staufen1 also reduced the abundance of XBP1 mRNA in unstressed cells, signifying that STAU1 is additionally required for the maintenance of XBP1 mRNA, and as such the sensitivity to ER stress. This poses the possibility that STAU1 promotes the re-ligation by RTCB after cleavage of XBP1 mRNA at a single site. Interestingly, while the endonuclease that catalyzes the cleavage step of XBP1 mRNA splicing is conserved in eukaryotes, RTCB catalyzes the ligation step of XBP1 mRNA splicing only in metazoans, co-occurring with the emergence of Staufen proteins. Taken together, the capacity of STAU1 to bind to different dsRNA sequences and structures has enabled the convergent evolution of SINE-directed Staufen-mediated mRNA decay in human and mouse, promotes the expression of IRAlus-containing mRNAs and the cellular resilience to ER stress.