SummaryA key feature of Notch signaling is that it directs immediate changes in transcription via the DNA-binding factor CSL, switching it from repression to activation. How Notch generates both a sensitive and accurate response—in the absence of any amplification step—remains to be elucidated. To address this question, we developed real-time analysis of CSL dynamics including single-molecule tracking in vivo. In Notch-OFF nuclei, a small proportion of CSL molecules transiently binds DNA, while in Notch-ON conditions CSL recruitment increases dramatically at target loci, where complexes have longer dwell times conferred by the Notch co-activator Mastermind. Surprisingly, recruitment of CSL-related corepressors also increases in Notch-ON conditions, revealing that Notch induces cooperative or “assisted” loading by promoting local increase in chromatin accessibility. Thus, in vivo Notch activity triggers changes in CSL dwell times and chromatin accessibility, which we propose confer sensitivity to small input changes and facilitate timely shut-down.

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The retromer-associated DNAJ protein Rme-8 is necessary for normal Notch recycling, and reductions in Rme-8 sensitize cells so that additional loss-of-sorting retromer or ESCRT-0 components have catastrophic effects.

Coordinating exit from the cell cycle with differentiation is critical for proper development and tissue homeostasis. Failure to do so can lead to aberrant organogenesis and tumorigenesis. However, little is known about the developmental signals that regulate the cell cycle exit-to-differentiation switch. Signals downstream of two key developmental pathways, Notch and Salvador-Warts-Hippo (SWH), and of myosin activity regulate this switch during the development of the follicle cell epithelium of the Drosophila ovary. Here, we have identified a fourth player, the integrin signaling pathway. We find that elimination of integrin function blocks mitosis-to-endocycle switch and differentiation in posterior follicle cells (PFCs), via regulation of the CDK inhibitor dacapo. In addition, we show that integrin mutant PFCs show defective Notch signalling and endocytosis. Furthermore, integrins act in PFCs by modulating the activity of the Notch pathway, as reducing the amount of Hairless, the major antagonist of Notch, or misexpressing Notch intracellular domain rescues the cell cycle and differentiation defects. Altogether, our findings reveal a direct involvement of integrin signalling on the spatial and temporal regulation of epithelial cell differentiation during development.

In Figure S3 J, the images of single FRAP examples were inadvertently removed during the production process. Below is the corrected version of the figure. The supplemental material PDF has been corrected. JCB: Correction