RNA- and Chromatin-binding Proteins in Small RNA-mediated Gene Silencing

[摘要] Gene expression is coordinately regulated at many levels to ensure the proper development and reproductive success of an organism. For example, RNA-binding proteins (RBPs) and microRNAs (miRNAs) mediate post-transcriptional gene silencing through recognition of target mRNA 3’ untranslated regions (UTRs). Pumilio/fem-3 binding factor (PUF) proteins are conserved from yeast to humans, where they bind PUF recognition elements (PREs) in mRNA 3’ UTRs to direct translational repression, mRNA storage, and transcript decay. Recent studies suggest that PUF proteins mediate more complex mechanisms of 3’UTR regulation. C. elegans puf-9 genetically interacts with the let-7 miRNA to regulate epidermal stem cell differentiation and vulval integrity at the larval-to-adult transition. However, the full extent of puf-9/miRNA interaction has not been explored. Here, we globally identify PUF-9 and miRNA binding sites in vivo, revealing that PUF-9 binding sites are enriched in 3’UTR regions with secondary structure, overlapping with miRNA binding sites. We show that the genetic interaction between puf-9 and let-7 is mediated by co-targeting of the lin-41 3’UTR. Altogether, our data are consistent with a model where PUF-9 and miRNAs bind to adjacent sites in structured 3’UTRs to co-regulate shared targets.In animals, germ cells are the only lineage that must be completely totipotent and indefinitely replenished at every generation. Together, small RNAs, histone modifying enzymes, and chromatin binding proteins control gene expression patterns required for germline function. The germline hereditary RNA interference (HRDE) pathway directs the establishment and maintenance of silenced heterochromatin. HRDE-1 Argonaute/small RNA complexes are passed down across generations to protect the immortal germ cell lineage. Here, we characterize the role of conserved chromatin binding protein MicrORChidia-1 (MORC-1) downstream of C. elegans siRNAs. We show that morc-1 is required for small RNA-guided nuclear silencing of operon pre-mRNAs. In addition, morc-1 is required for inheritance of RNAi across generations, but not for the biogenesis or inheritance of siRNAs themselves. Similar to hrde-1 Argonaute mutants, morc-1 mutants exhibit progressive sterility over multiple generations at elevated temperature and also mislocalize heterochromatin transgene reporters. Altogether, MORC-1 is likely an effector protein downstream of nuclear siRNA targeting.

[发布机构] University of Michigan

[效力级别] RNA binding protein

[关键词] small RNA;RNA binding protein;puf-9;morc-1;germline mortality;Genetics;Science;Human Genetics PhD