External Seminar Dealing with replication fork impediments, to break or not to break Seminar details May 23, 2017, 2:00 pm @ Small Lecture TheatreProf Marcel Tijsterman, Leiden University Host: Anton Gartner Abstract Faithful DNA replication is vital to prevent disease-causing mutations, chromosomal aberrations and malignant transformation. Accuracy, however, conflicts with pace and flexibility, and cells rely on specialized polymerases and helicases to ensure effective and timely replication of genomes that contain DNA lesions or secondary structures. How cells deal with permanent barriers to replication is unknown. We have recently shown that a single unresolved G-quadruplexed DNA structure can persist through multiple mitotic divisions without changing conformation. Failed replication across a G-quadruplex causes single-strand DNA gaps that give rise to DNA double-strand breaks in subsequent cell divisions, which are processed by polymerase theta (POLQ)-mediated alternative end-joining. Our data demonstrate that a single lesion can cause multiple unique genomic rearrangements, and that alternative end-joining enables cells to proliferate in the presence of mitotically inherited replication blocks. We have also shown that this POLQ-mediated alternative end-joining pathway is a major driver of inheritable genomic alterations in C. elegans. Unlike somatic cells, which employ non-homologous end joining (NHEJ) to repair DSBs, germ cells use polymerase theta in a conceptually simple mechanism that requires only one nucleotide as a template for repair. Also CRISPR/Cas9-induced genomic changes are exclusively generated through polymerase theta-mediated end joining, refuting a previously assumed requirement for NHEJ in their formation. Finally, through whole genome sequencing of propagated populations, we have shown that only POLQ proficient animals accumulate genomic scars that are abundantly present in genomes of wild C. elegans, pointing towards POLQ as a major driver of genome diversification. New mechanistic insights into how Pol Theta mediated end joining (TMEJ) processes DNA breaks in mammalian cells will be discussed.