Cell cycle regulation of differentiation in development and disease
Dept of Oncology and Wellcome/MRC Cambridge Stem Cell Institute, University of Cambridge.
It is essential that division and differentiation are carefully co-ordinated during development. Moreover, a disruption of this co-ordination is a central characteristic of many forms of cancer. We are investigating the molecular mechanisms by which the cell cycle machinery can directly control the activity of key transcription activators, the proneural proteins, which are responsible for regulating differentiation in the nervous system, pancreas, gut and multiple other tissues. We see that chromatin binding and transcriptional activity of basic helix-loop-helix transcription factors of the "proneural" family, which regulate fate choice and differentiation in many tissues, is regulated by multi-site phosphorylation by cell cycle kinases. Inhibiting proneural protein phosphorylation preferentially activates targets associated with driving cell cycle exit and differentiation, and this is key to altering the balance away from maintenance of a proliferative state. Our results lead us to a model whereby proneural transcription factor post-translational modification intersects directly with the epigenetic landscape of downstream targets to determine whether cells maintain their progenitor status or undergo differentiation during development and in adult homeostasis in multiple tissues. For instance supporting this model using engineered, we see that post-translational modification of proneural proteins controls both the formation of endocrine cells in the developing pancreas and the ability of intestinal secretory progenitor cells to respond to tissue injury by contributing to repair. We are also investigating how proneural protein phosphorylation by sustained cdk activity may result in a fundamental shift in the behavior of cancer cells, disabling their ability to undergo mitotic arrest and terminal differentiation. In particular, the paediatric cancer neuroblastoma is associated with stalling of normal differentiation resulting in excessive proliferation of neuroblastic precursors of the sympathetic nervous system. We see that CDK-dependent phosphorylation of the proneural protein ASCL1 in in neuroblastoma plays a critical role in controlling the balance between cell proliferation and differentiation; preventing CDK-dependent phosphorylation of ASCL1 results in changes in the genome-wide transcriptional programme of neuroblastoma cells, leading to suppression of pro-proliferative targets and simultaneous activation of genes that drive cell cycle exit and differentiation. Mechanistically, ASCL1 ChIPSeq reveals that dephosphorylation of Ascl1 leads to a disabling of the super-enhancer network that supports the progenitor state and enhanced binding of un(der)phosphorylated ASCL1 at sites associated with pro-differentiation targets. Finally, we also show that chemical CDK inhibition is sufficient to drive differentiation of neuroblastoma cells in a manner dependent on endogenous ASCL1. Therefore, we conclude that CDK-dependent phosphorylation of ASCL1 acts as a critical fulcrum controlling the balance between proliferation and differentiation and thus points to novel therapies for neuroblastoma. Furthermore, post-translational control of may play a key role in regulation of other proneural proteins that have been shown to act as lineage-specific oncogenes in a number of settings.