mRNP Granules: Novel Insights Into Assembly, Composition and Relevance to Disease

Abstract

Post-transcriptional processes are crucial in the regulation of gene expression. Messenger ribonucleoprotein (mRNP) granules are dynamic, self-assembling membraneless organelles that harbor non-translating mRNAs implicated in multiple post-transcriptional processes like mRNA translation, repression, localization and turnover. Besides their involvement in cytoplasmic mRNA biology, mRNP granules are also associated with disease such as neurodegenerative diseases and cancer. In this thesis, we focus on P-bodies (PBs) and stress granules (SGs) studying their assembly, composition and relevance to disease. In Chapter 2, we studied PB assembly and found that a specific mRNA RPS28B is important for P-body assembly by acting as a scaffold that enhances the interaction between Edc3, an important PB assembly protein recruited to the RPS28B 3’UTR, with Rps28 protein being translated off of the mRNA. The Edc3-Rps28 interaction correlates with PB assembly. Our work suggests that PBs may be preferentially nucleated by specific mRNA scaffolds, possibly a common theme in mRNP granule assembly. Furthermore, this is the first description, in yeast, of a cis-translated protein interacting with a protein recruited to the 3’UTR of the same mRNA, which in turn has functional consequences for assembly of cellular structures. In Chapter 3, we studied the composition of SGs with the long term goal of uncovering additional functions of SGs. We developed novel SG purification protocols that are more accurate and sensitive in identifying SG components than previously published protocols. We have identified novel SG proteins that suggest that SGs could have roles in modulating translation during stress by altering the cellular tRNA charging status and cell cycle progression by sequestering Cdc28/CDK kinases, an area of immediate future interest. In Chapter 4, we studied the relevance of SGs to neurodegenerative disease. We examined the claims that SGs are important for TDP-43 aggregation and toxicity associated with ALS. We looked at SG mutants and overexpression of SG proteins with regards to TDP-43 toxicity and aggregate formation in an established yeast model and in mammalian cell lines and found that SG assembly facilitates but is not required for TDP43 aggregation.