Encoding Cell Cycle Regulatory Information in the Genome

Abstract

The timing of cell cycle exit and initiation of terminal differentiation must be precisely coordinated to ensure proper development of many tissues. Additional cell cycles during development can disrupt, but not necessarily prevent, progression of terminal differentiation programs, leading to tissues with incorrect cell numbers and morphology. The mechanisms that coordinate the transition from a proliferative state to a fully differentiated post-mitotic state are not well understood, and even less is understood about how a non-cycling postmitotic state is maintained in terminally differentiated cells. The eyes and wings of the fruit fly Drosophila melanogaster are excellent tissues to study this phenomenon, as both tissues undergo a relatively synchronous final cell cycle before exiting the cell cycle permanently at 24h after the start of metamorphosis, coincident with visible progression of terminal differentiation programs. Cell cycle exit in Drosophila wings and eyes involves the transcriptional silencing of hundreds of cell cycle genes. However, maintaining cell cycle exit relies on preventing the re-activation of three rate-limiting cell cycle genes, the G1-S cyclin, Cyclin E, the cell cycle transcriptional activator, E2F1 and the regulator of mitotic entry, cdc25c, termed String in flies. Our prior work established that after cell cycle exit, chromatin accessibility is reduced at potential regulatory elements for these three genes, leading to a hypothesis that closing chromatin maintains cell cycle exit by preventing activation of the rate-limiting cell cycle genes. In this thesis I examine this hypothesis by developing new techniques to allow for more detailed assays of chromatin accessibility changes and chromatin modifications (Chapter 2). I also test and validate several regulatory elements for the e2f1 and string loci, to determine which elements are tissue specific vs. shared for the wing and eye and examine their shut off dynamics during chromatin accessibility changes after cell cycle exit (Chapter 3). Finally, I identify a chromatin remodeler responsible for the closing of chromatin accessibility at the string locus, and determine that it works together with a transcription factor expressed during metamorphosis to coordinate chromatin accessibility changes that decommission enhancers at cell cycle genes and early differentiation genes, to maintain cell cycle exit as terminal differentiation progresses (Chapter 4). Altogether this work examines how complex cell cycle regulatory events can be encoded in the genome, to ensure the proper coordination of cell cycle control with cellular differentiation.