Department of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati College of Medicine, Cincinnati, OH, USA.
Institute of Biochemistry, University of Giessen, Giessen, Germany.
Cell Rep. 2019 Jan 22;26(4):845-854.e6. doi: 10.1016/j.celrep.2018.12.097.
Notch is a conserved signaling pathway that is essential for metazoan development and homeostasis; dysregulated signaling underlies the pathophysiology of numerous human diseases. Receptor-ligand interactions result in gene expression changes, which are regulated by the transcription factor RBPJ. RBPJ forms a complex with the intracellular domain of the Notch receptor and the coactivator Mastermind to activate transcription, but it can also function as a repressor by interacting with corepressor proteins. Here, we determine the structure of RBPJ bound to the corepressor SHARP and DNA, revealing its mode of binding to RBPJ. We tested structure-based mutants in biophysical and biochemical-cellular assays to characterize the role of RBPJ as a repressor, clearly demonstrating that RBPJ mutants deficient for SHARP binding are incapable of repressing transcription of genes responsive to Notch signaling in cells. Altogether, our structure-function studies provide significant insights into the repressor function of RBPJ.
Notch 是一条保守的信号通路，对后生动物的发育和体内平衡至关重要；信号失调是许多人类疾病病理生理学的基础。受体-配体相互作用导致基因表达的变化，这些变化受转录因子 RBPJ 的调控。RBPJ 与 Notch 受体的细胞内域和共激活因子 Mastermind 形成复合物，以激活转录，但它也可以通过与核心抑制蛋白相互作用而发挥抑制作用。在这里，我们确定了与核心抑制蛋白 SHARP 和 DNA 结合的 RBPJ 的结构，揭示了其与 RBPJ 结合的模式。我们在生物物理和生化细胞测定中测试了基于结构的突变体，以表征 RBPJ 作为抑制剂的作用，清楚地表明，与 SHARP 结合缺陷的 RBPJ 突变体不能抑制对 Notch 信号有反应的基因在细胞中的转录。总的来说，我们的结构-功能研究为 RBPJ 的抑制功能提供了重要的见解。
