Structural principles governing kinase specificity towards RNA polymerase II C-terminal domain and their impact on transcription

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2023-05

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Transcription, one of the major processes in the cell, is the conversion of genetic information (DNA) into a temporarily stored information (RNA). To orchestrate, selectively regulate and control this conversion, cells have evolved a variety of tools – the most interesting being the CTD of Pol II. The CTD sequence consists of seven amino acids repeated multiple times (Up to twenty-six repeats in yeast and fifty-two repeats in humans). Phosphorylation is just one of the many post-translational modifications carried out on the CTD, however it is arguably the most important for transcriptional regulation since five of the seven residues on every consensus heptad are capable of phosphorylation. In Chapter 2, I elucidate how kinases can correctly recognize the right residue for phosphorylation. Data presented in this chapter reveals the underlying rules of substrate recognition by CTD kinases which is important for the timely regulation of transcription. Furthermore, in Chapter 3, I describe the experimental setup and methodology utilized to study the spatiotemporal pattern of phosphorylation across a full-length yeast CTD substrate. We used this setup to identify the pattern of phosphorylation conducted by CDK7 and to identify its capability of selectively preferring heptads. Utilizing the knowledge of the specificity of kinases, we use them to identify interactors by generating phosphorylated substrates in vitro and identifying differential interactors as explained in Chapter 4.

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