Genomic instability is at the root of several human diseases, such as cancer and neurological disorders. Therefore, understanding the pathways that counteract it is pivotal to directing a long-term strategy to improve human health. A recognised source of genomic instability is the interference between two processes that are essential to life and share the same substrate, the chromatin: transcription and DNA replication. Head-to-head collisions between the replication fork and RNA polymerase may result in the formation of DNA-RNA triple helices, R-loops, DNA damage and genomic instability. How the conflicts between transcription and DNA replication are avoided or dealt with is a fundamentally important biological question, still largely unanswered.
Work from my group suggests that separating in space and time these two processes could help reducing transcription-replication conflicts. We suggest that the spatio-temporal organisation of DNA replication ensured by the DNA replication-timing program could be key to the coordination of DNA replication and transcription.
My group has discovered the first mammalian genome-wide regulator of replication timing, RIF1, also demonstrating that it coordinates the temporal and spatial aspects of DNA replication. This project will focus on understanding the molecular basis of the RIF1-dependent spatial segregation of early- and late-replicating genomic regions.
We have discovered that the majority of RIF1 associates with large chromatin domains (RIF1-associated domains, or RADs), coating the late-replicating genome. We propose that the assembly of the large RIF1 arrays forming the RADs is essential for the RIF1-dependent functional organisation of chromatin and we aim to understand:
1. How does RIF1 assemble into RADs?
2. How are the RADs segregated to the nuclear periphery?
To address these questions, we will employ mouse embryonic stem cells as a model system, and a wide range of techniques, including Cas9/CRISPR-mediated genome engineering, immunoprecipitation, chromatin immunoprecipitation, in vitro pull downs and imaging, including super-resolution microscopy, FRET and FRAP. We are looking for enthusiastic, flexible and hard-working candidates driven by curiosity and passion for science. Basic knowledge of cell/molecular biology and ability to work in English, in an international environment are required. The student will lead the project and work with the support of excellent, state-of-the-art facilities and well-experienced postdoc and students in the lab. Direct supervision from the principal investigator and the colleagues in the lab, together with access to relevant courses will help developing all the necessary skills to successfully complete the project.
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