Role of DNA replication timing in gene expression and chromatin organisation

Eukaryotic DNA is duplicated according to an evolutionary conserved temporal pattern. This pattern of DNA replication is altered during development and differentiation and can be dysregulated in cancers. While temporal changes in genome duplication are associated with altered transcription and chromatin organisation, it is still unknown whether DNA replication timing (RT) is a cause or a consequence of cellular fate changes. During my thesis I used a conditional system to perturb DNA RT in a single cell cycle in budding yeast, in combination with whole-genome sequencing techniques such as replication profiles, RNA-Seq and MNase-Seq in order to understand the biological importance of a defined pattern of genome replication and the impact on the genome structure and function. Overall, dramatic changes in gene expression, chromatin structure and transcription-factor (TF) binding events were observed, and a significant number of genes affected are involved in differentiation processes such as sporulation. While some differentially expressed genes showed significant chromatin changes, there were also examples where this was not the case, as well as genes with changes in chromatin and no changes in expression, which illustrates the complex nature of the relationship between RT, gene expression and chromatin. Differential TF binding events explained some of the observed changes, supporting a role for RT to maintain the correct TF binding dynamics during S-phase. Additionally, the fact that budding yeast origins are defined by specific sequences allowed the local modulation of RT which showed a direct effect of RT on gene expression. Altogether, the work generated during this thesis provides insight into the complex relationship between replication timing, gene expression and the chromatin landscape.