Cell Reports
Volume 26, Issue 4, 22 January 2019, Pages 845-854.e6
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Structural and Functional Studies of the RBPJ-SHARP Complex Reveal a Conserved Corepressor Binding Site

https://doi.org/10.1016/j.celrep.2018.12.097Get rights and content
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Highlights

  • The corepressor SHARP binds the transcription factor RBPJ in a bipartite manner

  • SHARP binds the BTD and CTD of RBPJ, using motifs analogous to other corepressors

  • Structure-based mutants affect RBPJ-SHARP complex formation in vitro and in cells

  • RBPJ mutants are defective in repression from Notch target genes in cells

Summary

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.

Keywords

Notch signaling
CSL
RBPJ
SHARP
MINT
SPEN
transcriptional regulation
signal transduction
X-ray crystallography
isothermal titration calorimetry

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