Developmental control of cell cycle exit in Drosophila
Bruce Edgar
Fred Hutchinson Cancer Research Center, Seattle, USA
Terminal differentiation is frequently coupled with permanent exit from the cell cycle, yet it remains unclear how cell proliferation is blocked in differentiated tissues. We have examined this process in Drosophila wings and eyes and discovered that cell cycle exit involves a “double assurance” mechanism in which the activities of the transcription factor, E2F, and a G1 Cyclin/CDK complex, CycE/Cdk2, are independently and dominantly silenced. Bypassing this mechanism by co-expression of E2F and CycE promotes the indefinite proliferation of terminally differentiated cells in vivo. In some differentiating cell types (e.g. neurons) known cell cycle inhibitors including the retinoblastoma homolog, Rbf, and the p21/p27 homolog, Dacapo, contribute to parallel repression of E2F and Cyclin E/Cdk2. In other cell types however (e.g. wing epithelial cells), additional unknown mechanisms act to inhibit E2F and Cyclin/Cdk activity to enforce permanent cell cycle exit upon terminal differentiation. Since the critical targets of both E2F and Cdk2 reside on chromatin at transcriptional promoters and origins of replication, we like the idea that differentiating cells may express factors that shield these targets from E2F binding and Cdk2 phosphorylation. To understand cell cycle exit and find the factors that mediate it, we are performing candidate gene tests, gene expression profiling, and forward genetic screens. Our genetic screens employ an E2F-responsive pcna-white+ reporter gene to identify, using eye color, loss- and gain-of-function mutations that cause perdurant E2F activity in the differentiating Drosophila eye. Secondary screens then identify those genes that can drive ectopic cell proliferation. To date we’ve screened ~180,000 mutant chromosomes covering ~60% of the genome, and ~4000 gain-of-function “EP” transgenics. We’ve identified 5 mutant loci and ~10 over-expressed genes that deregulate E2F activity and drive unscheduled cell proliferation in differentiating wings and eyes. Three genes were expected (E2F, CycE, Ago/Fbw7), but most are genes not previously known to be involved in cell cycle control. Some of these novel cell cycle regulators, and our working model for cell cycle exit, will be discussed.
