Transcription pre-initiation complex regulation by nutrient dependent O-GlcNAcylation of TATA-Box Binding Protein – TaNGoT

Gene expression is performed by three RNA polymerases (RNAPs) each of which is devoted to a specific class of gene. Assembly of specific molecular complexes at gene promoters to form pre-initiation complexes (PICs) ensure recruitment and positioning of the correct RNAP. Among the variety of factors necessary to form a functional PIC, the TATA-Box Binding Protein (TBP) has been considered as a common universally conserved platform for PIC assembly with a passive role in gene expression regulation. However, our recent data showed that TBP is a key player in the dynamic regulation of transcription initiation and that it is dynamically modified by O-GlcNAcylation at T114, T126 and S158. O-GlcNAcylation is a finely tuned post-translational modification of nucleocytoplasmic protein controlled by two unique enzymes. The O-GlcNAc transferase adds the sugar whereas O-GlcNAcase removes it. O-GlcNAcylation dynamically regulates activities of molecular players virtually involved in all cellular processes ranging from cell signaling, metabolism, epigenetics and transcription. It is now clear that O-GlcNAcylation plays critical roles in nutrient and stress regulation of nearly all aspects of cellular physiology. A growing body of evidence places O-GlcNAcylation at the nexus of nutritional status, cellular homeostasis, and the establishment of chronic diseases beyond those typically referred to as metabolic-related pathologies.
We previously showed that TBP T114-O-GlcNAcylation is involved in the formation of the B-TFIID complex, hence the dynamic association of TBP to chromatin and TBP promoter redistribution. In terms of cellular physiology, the glycosylation of TBP at T114 regulates a set of genes involved in lipid metabolism. Yet, the primary roles of T126 and S158 O-GlcNAcylation remain unknown but our preliminary data suggest that they could potentially be involved in the regulation/assembly of the PICs requested for transcription initiation by RNAPI, RNAPII and/or RNAPIII. Literature shows that alternative assembly of PICs allows targeting of specific core promoter elements, transcription of a subset of genes, and dynamism of transcription initiation by RNAPs. Therefore, site-specific O-GlcNAcylation of TBP may have a profound impact on cell transcriptome and cell physiology. Here, I propose to document the site-specific function of these residues, using combined transcriptomic and proteomic approaches on CRISPR-edited cell lines along with structure conformation and DNA-bending activity studies. Phenotypes of the CRISPR-edited cell lines will be further investigated to link the role of each O-GlcNAcylation site of TBP on cell physiology to the underlying molecular mechanism. Overall, TaNGoT ambitions to 1) understand the role of the TBP regulation by O-GlcNAcylation on transcription initiation and its impact on cell physiology, 2) decipher the O-GlcNAcylation site-specific functions onto the assembly of the PICs, 3) unveil the intrinsic molecular mechanism of this regulation. The outcome of TaNGoT will update the dogma of TBP as a mere platform by describing the molecular mechanism of regulation of basal transcription machinery by O-GlcNAcylation of TBP, highlighting an “O-GlcNAc code” of TBP.