Elucidating the propagation and function of H2A and H2B modifications across DNA replication

ended

Chromatin and its accompanying histone marks are fundamental for correct, cell type-specific gene expression and thus important to ensure cellular identity of each cell. Importantly, this requires that the chromatin landscape is accurately propagated and maintained across DNA replication, a process which is still not well understood. While there have been insights into how histone H3 and H4 marks are transmitted to daughter cells, information on H2A and H2B marks are still missing. Using mouse embryonic stem cells, I will gain first insights into how modifications on H2A and H2B are maintained during DNA replication, and their impact on restoring the parental chromatin landscape on replicated DNA, with the following three specific objectives: (1) I will investigate the propagation principles and kinetics of H2A/H2B modifications during DNA replication using a combination of advanced mass spectrometry and genomic approaches with a focus on ubiquitinated H2A/H2B. (2) I will gain mechanistic insights into the importance of H2A/H2B ubiquitination on chromatin reestablishment with a degron-based depletion approach targeting H2A/H2B ubiquitin ligases. This will allow me to study if, where and how the H2Aub and H2Bub-dependent histone marks are reestablished and hence reveal important features of histone mark crosstalk during DNA replication. (3) I will determine whether H2A-H2B dimers are propagated symmetrically to the two daughter strands by employing a strand-specific sequencing approach, and test mutants wherein symmetric H3-H4 propagation is compromised. This proposal will provide pioneering insights into how histones H2A and H2B are propagated and unveil the epigenetic inheritance of their marks, and also bring seminal understanding of histone mark crosstalk during DNA replication and across cell division. This will open new research avenues in chromatin research and give me an unique opportunity to follow up on these mechanisms as future, independent researcher.