Investigating RNP granules as centres of novel translational regulation in eukaryotic cells

Translation of mRNA to protein and its attendant regulation underpins all life since proteins are the principal effectors of biological function; they are responsible for catalysing most biochemical reactions as well as many structural and regulatory roles. Although translation is a generally well understood process common to all eukaryotes (animals, plants and fungi) it is becoming increasingly clear that the localisation of mRNA within the cell is also critically important and can play roles in the degradation, storage and the translation process itself. This can occur within so-called Ribonucleoprotein-containing bodies or granules, which we term here RNP granules, of various different types and physical properties ranging from aggregates to 'liquid droplets'. These granules are often linked to 'stress' conditions and can have different functions, as well as being linked to a number of neurodegenerative and musculodegenerative diseases such as Fragile X mental retardation, spinal muscular atrophy, Huntington's and Alzheimer's.
Our hypothesis is that there are a number of related RNP granules with overlapping as well as specific mRNA/protein components that together engender distinct functionality. To address this we will comprehensively characterize the dynamic membership of these granules in yeast under a number of specific stress conditions, focussing on functionality i.e. the fate of mRNAs in different granules. Hence, we will use RNA-immunoprecipitation and RNA-seq, coupled with quantitative proteomics and integrative bioinformatics to comprehensively characterise the granular composition of 4 RNP types (stress granules, P-bodies, mRNA granules and protein aggregates). This will be done under different stresses/levels, in order to generate the granules and observed how different mRNA and protein components change over time. We will validate our findings for candidate mRNAs and proteins using cutting-edge immunofluorescence and imaging techniques. This work builds on our collective efforts to map and understand how eukaryotic cells regulate the translation of their specific mRNAs into proteins in our labs, where this occurs, and which proteins are involved including under stresses such as heat shock.
The project will combine wet-lab biology: protein biochemistry, molecular biology, and yeast genetics (core Bioscience skills) informed by cutting-edge next-gen RNA sequencing, quantitative proteomes and computational biology. This combination of lab and computational approaches will allow a 'systems' approach to understand the biology, with one feeding into the other to exploit the so-called 'new way of working'. The project will fully characterise novel cellular bodies containing proteins and associated mRNAs that can regulate protein synthesis in times of cell stress. This is fundamental bioscience that will generate tools applicable to the optimal expression of recombinant proteins from yeast cell cultures.