Fechner Johannes, Ketelhut Manuela, Maier Dieter, Preiss Anette, Nagel Anja C
Department of General Genetics 190g, University of Hohenheim, Garbenstr. 30, 70599 Stuttgart, Germany.
Int J Mol Sci. 2022 Oct 15;23(20):12336. doi: 10.3390/ijms232012336.
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 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.
Notch的主要作用是确定细胞身份，借此细胞对Notch信号活性中极其微小的变化做出反应。因此，Notch组分的剂量对于调节至关重要。Notch信号转导的核心是CSL蛋白：它们与各自的辅因子一起介导Notch靶基因的激活或沉默。CSL蛋白在物种间的序列和结构极为相似。我们注意到果蝇同源物无翅抑制因子（Su(H)）在转录复合物中是稳定的。利用特定的转基因果蝇品系和HeLa RBPJ细胞，我们提供证据表明，如果不通过与共抑制因子无翅或共激活因子Notch结合来保护，Su(H)会经历蛋白酶体降解，半衰期约为两小时。此外，Su(H)的稳定性受MAPK依赖性磷酸化控制，这与人类细胞中RBPJ的早期数据相符。然而，同源的鼠类和人类RBPJ蛋白在我们的系统中对降解具有很大抗性。然而，突变假定的蛋白质接触位点会使RBPJ对蛋白水解敏感。总体而言，我们的数据突出了跨物种CSL蛋白稳定性调节的相似性，并暗示CSL蛋白的周转可能是在转录水平直接调节Notch信号输出的一种保守方式。
