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Comprehensive analysis of the proximity-dependent nuclear interactome for the oncoprotein NOTCH1 in live cells.
Journal of Biological Chemistry 2023 December 2
Notch signaling plays a critical role in cell fate decisions in all cell types. Furthermore, gain-of-function mutations in NOTCH1 have been uncovered in many human cancers. Disruption of Notch signaling has recently emerged as an attractive disease treatment strategy. However, the nuclear interaction landscape of the oncoprotein NOTCH1 remains largely unexplored. We therefore employed here a proximity-dependent biotin identification (BioID) approach to identify in vivo protein associations with the nuclear Notch1 intracellular domain (NICD) in live cells. We identified a large set of previously reported and unreported proteins that associate with NOTCH1, including general transcription and elongation factors, DNA repair and replication factors, coactivators, corepressors, and components of the NuRD and SWI/SNF chromatin remodeling complexes. We also found that NICD associates with protein modifiers and components of other signaling pathways that may influence Notch signal transduction and protein stability such as USP7. We further validated the interaction of NOTCH1 with HDAC1 or GATAD2B using protein network analysis, proximity-based ligation, in vivo crosslinking and co-immunoprecipitation assays in several Notch-addicted cancer cell lines. Through data mining, we also revealed potential drug targets for the inhibition of Notch signaling. Collectively, these results provide a valuable resource to uncover the mechanisms that fine-tune Notch signaling in tumorigenesis and inform therapeutic targets for Notch-addicted tumors.
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