The Binding of CSL Proteins to Either Co-Activators or Co-Repressors Protects from Proteasomal Degradation Induced by MAPK-Dependent Phosphorylation

The primary role of Notch is to specify cellular identities, whereby the cells respond to amazingly small changes in Notch signalling activity. Hence, dosage of Notch components is crucial to regulation. Central to Notch signal transduction are CSL proteins: together with respective cofactors, they mediate the activation or the silencing of Notch target genes. CSL proteins are extremely similar amongst species regarding sequence and structure. We noticed that the fly homologue suppressor of hairless (Su(H)) is stabilised in transcription complexes. Using specific transgenic fly lines and HeLa RBPJ(KO) cells we provide evidence that Su(H) is subjected to proteasomal degradation with a half-life of about two hours if not protected by binding to co-repressor hairless or co-activator Notch. Moreover, Su(H) stability is controlled by MAPK-dependent phosphorylation, matching earlier data for RBPJ in human cells. The homologous murine and human RBPJ proteins, however, are largely resistant to degradation in our system. Mutating presumptive protein contact sites, however, sensitised RBPJ for proteolysis. Overall, our data highlight the similarities in the regulation of CSL protein stability across species and imply that turnover of CSL proteins may be a conserved means of regulating Notch signalling output directly at the level of transcription.

Automated summary

The primary role of Notch is to specify cellular identities by responding to small changes in Notch signaling activity. CSL proteins play a central role in Notch signal transduction and their stability is regulated similarly across species. The study shows that the fly homologue Su(H) is stabilized in transcription complexes and its stability is controlled by MAPK-dependent phosphorylation and binding to other proteins. These findings highlight the conservation of CSL protein stability in regulating Notch signaling output at the transcription level.