Mechanistic Insights into Coactivator Recruitment by Nuclear ReceptorsPublic
Nuclear receptors (NRs) are an important family of transcription factors that often regulate genes in response to ligands and by way of direct interactions with coactivator proteins. Many NR-coactivator pairs have been identified that cooperate to regulate transcription but fully understanding how NRs recruit specific coactivators involves learning which of their domains interact and how the respective structural and biochemical features govern their associations. Here, I investigated the conformation, dynamics, and interactions of NR and coactivator domains, with the goal of gaining deeper insights into molecular functions. Initially, I characterized the ligand-binding domain (LBD) of Drosophila Ftz-F1 receptor, which is hypothesized to recruit coactivators in a ligand-independent manner. My studies showed that the canonical ligand-binding pocket, which appeared occupied by a short segment of the protein in the crystal structure, is dynamic, and likely samples an open conformation in solution, suggesting that Ftz-F1’s recruitment of coactivators could be modulated by ligand binding. I then investigated how another atypical NR, Nurr1, which was also thought to recruit coactivators in a ligand-independent manner, can recruit the coactivator SRC1 in an LBD-independent manner. Specifically, my studies show that the Nurr1 activation function-1 (AF1) domain is unstructured in solution and contains a ~20-residue segment that binds to the SRC1 PAS-B domain, which represents a novel mechanism for coactivator recruitment by NRs. Additionally, structure-function analysis of the interaction revealed that AF1 likely forms a helix upon binding and engages with the same surface on PAS-B as the transcription factor STAT6, suggesting that binding this surface represents a conserved mechanism for transcription factors to recruit SRC1. Since PAS domains are known to bind small molecules, I then investigated whether the SRC1 PAS-B domain could bind small molecule ligands. My studies showed that a conserved, but previously uncharacterized, pocket can bind small molecules such as prostaglandins, suggesting a potentially novel mechanism for regulating interactions between PAS-B and transcription factors. Collectively, my results have broadened our understanding of how dynamics, ligand-binding, and structure affect NR-coactivator interactions and, in the process, have uncovered new mechanisms of transcriptional activation.
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