Regulation of Protein Production Dynamics: RNA Binding Proteins and the Ribosome Code.
Gene expression in mammalian cells is tightly controlled at several levels - from transcription to translation and protein degradation. However, to date, most studies focused on changes in overall mRNA abundance. Yet, the mammalian genome encodes over 1,500 RNA binding proteins (RBPs), several of which are recurrently mutated in diseases, such as cancer and neurological disorders, suggesting that post-transcriptional gene expression regulation and especially mRNA translation are important in both health and human disease. Furthermore, the regulatory role of the ribosome itself has so far been under-explored. Evidence is mounting that specialized ribosomes, which vary in ribosomal protein stoichiometry and post-translational modifications (PTMs), exist that may impact the translation of specific mRNAs through an as-yet-undefined ‘ribosome code’.
Our goal is to understand the principles and mechanisms by which translational regulation controls the dynamics of gene expression and therefore affects processes like differentiation, stress response and pathogenesis. We are focusing on two specific aspects of translational control.
First, we systematically identify and characterize RBPs that regulate translational changes. We are combining unbiased, high-throughput CRISPR-based screening against all 1,500 predicted RBPs with global measurements of RNA dynamics, and protein production and degradation. This will link RBPs to their mRNA targets, providing the foundation for future detailed functional follow-ups, allowing us to elucidate functional and causal insights of how RBPs regulate mRNA translation.
Second, we are determining the extent of ribosomal heterogeneity by high accuracy mass spectrometry, focusing right now on two potential sources of ribosomal heterogeneity – differential expression in core ribosomal proteins (RPs) and changes in their post-translational modifications. Based on these measured changes in ribosome composition, we are selecting RPs and PTMs with the strongest changes for further functional characterization. The detailed follow up will provide for a selected set of RPs and PTMs the principles and mechanistic insight how ribosome specialization regulates translation.
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