Final Report Summary - RNPLOCALIZATION (Towards a mechanistic understanding of mRNA localization)
The subcellular localization of mRNAs is a critical mode of controlling gene expression during animal development and eukaryotic cell function. The goal of this project was to gain fundamental insights into the molecular basis of the cytoplasmic regulation of gene expression by mRNA localization. We used a combination of biochemical, structural and genetic approaches. During the project, we carried out a systematic biochemical and structural mapping of mRNA localization complexes to elucidate the molecular networks underlying the process. We also tested functional atomic-level predictions in vivo by focusing on early development in Drosophila. We investigated novel types of RNA-binding proteins as well as canonical ones. We determined the crystal structures of functional domains, protein-protein and protein-RNA complexes. We combined the information obtained with a thorough in vitro analysis of the features and RNA-binding properties of the proteins. For example, we used this multidisciplinary approach to characterize Exuperantia, a protein with a key function in the localization of mRNAs essential for development. We solved the structure of Exuperantia and studied its regulation and RNA-binding specificity. Our work provides a mechanistic explanation for the phenotype of classical fly mutants. In another project, we characterised the RNA-binding specificity and structure of Staufen, a conserved protein involved in mRNA localization in flies and humans. We discovered that Staufen directly recognize specific bases in stem-loop structures of its target mRNAs. We also carried out a large-scale protein-protein interaction screen and identified novel interactions and novel components interacting with known mRNA localization factors. Our combination of biochemical, structural and in vivo approaches allowed us to gain unique mechanistic details into mRNA localization, a conserved and essential process.