mRNA turnover is a key step in regulation of gene expression. Its mis-regulation has been linked to a range of human disorders. The long-term goal of my laboratory is to understand the mechanism and regulation of mRNA turnover in humans. Removal of the mRNA 5'cap by the process of decapping is an important step in mRNA turnover. Recent evidence has shown that the protein complex responsible for decapping co-localizes with other factors involved in degradation of mRNA from the 5'end in sub-cytoplasmic foci termed processing bodies. Moreover, compelling data shows that processing bodies are active in mRNA decay. This suggests that mRNA decay can be a localized process. Here we will test the role of human processing bodies in three specific mRNA decay processes that play a key role in human gene expression and disease: All-rich element (ARE)-mediated mRNA decay, nonsense-mediated decay (NMD) and mRNA decay triggered by micro (mi)RNAs. We will employ in situ hybridization, indirect immunofluorescence, co- immunoprecipitation and mRNA decay assays to test how these mRNA decay pathways interface with processing bodies. In addition, we will test if catalytic components of PBs are targeted for proteolysis when localized outside of PBs where they may stimulate promiscuous RNA decay. These experiments are aimed at addressing the following questions: i) How are unstable mRNAs targeted to processing bodies and ii) why are mRNA decay enzymes concentrated in PBs rather than throughout the cytoplasm? This proposal is relevant to public health because mis-regulation of the degradation of protein-encoding mRNAs has been linked to a range of human disorders. Understanding the mechanism behind mRNA turnover will help develop strategies to fight such diseases.
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