microRNA (miRNA)-directed translational repression is a process in which miRNA serves as a guide for an RNA induced silencing complex (RISC) to bind to homologous target mRNA and arrest translation. It is used to regulate gene expression and implicated in a variety of biological processes such as development and heterochromatin formation. Much research over the last decade has focused on the mechanisms underlying miRNA-directed translational control, but questions remain about the fate of mRNA following translational repression. The fate of repressed mRNA is important considering how many developmental and epigenetic disorders could be caused by regulatory defects in miRNA-directed translational repression. P bodies are known to be degradation sites for miRNA-regulated mRNA, but they may also be sites of mRNA storage. Stress granules have been implicated in storing mRNA under adverse conditions for future reactivation of translation. They could also be storage or degradation sites for miRNA-regulated mRNA. Additionally, recent studies demonstrate that it is possible to reactivate translation of miRNA-regulated mRNA localized to P bodies. 1 propose to simultaneously visualize and quantify mRNA, P body, and stress granule levels to track the fate of miRNA-regulated mRNA and design a biological tool to control miRNA- directed translational repression in order to determine under what conditions miRNA-regulated mRNAs are stored or degraded. I will use RNAi knockdowns of P body and stress granule components to determine which elements are responsible for deciding the fate of miRNA-regulated mRNA and to identify factors involved in reactivating miRNA-regulated mRNA for translation. The proposed work will establish the relationship between miRNA-directed translational repression and P body and stress granule components. Results from this work will serve as a foundation for more rational design of miRNA-based therapeutic tools for the treatment of human disease and genetic disorders. Additionally, as miRNA-directed translational repression affects many developmental and epigenetic processes, understanding the fate of miRNA-regulated mRNA will reveal how defects in miRNA-directed translational repression affect downstream processes in developmental and epigenetic disorders.
|Shih, Joseph D; Waks, Zeev; Kedersha, Nancy et al. (2011) Visualization of single mRNAs reveals temporal association of proteins with microRNA-regulated mRNA. Nucleic Acids Res 39:7740-9|