Polymerase theta-mediated end joining in telomere maintenance

Polymerase theta (POLQ) defines a novel error-prone pathway of DNA double-strand break (DSB) repair in which the outermost nucleotide of one end serves to prime DNA synthesis on the other end of the break. This provides a fast, simple and elegant mechanism of DNA repair and protection against genome instability. It is no surprise therefore that POLQ deficiency is able to highly sensitise cells to the lethal accumulation of DSBs induced by ionizing radiation and, conversely, that the tumours most resistant to radiotherapy have high levels of POLQ expression. Here, we propose to investigate whether POLQ could also mediate joining of uncapped telomeres, which at molecular level resemble DSBs introduced at a specific chromosomal location. Loss of chromosome capping occurs during physiological processes such as cellular senescence and ageing, due to insufficient retention of the telomere-repeat binding factor TRF2. The unprotected ends are re-joined with formation of chromosome end-to-end fusions and trigger the rampant genome instability that can drive tumorigenesis. In this project, we will uncap telomeres using shRNA-mediated inactivation of TRF2 and we will evaluate the impact of POLQ inhibition on their processing and the formation of telomeric fusions. As these fusions arise by classical or alternative non-homologous end joining (C-NHEJ or A-NHEJ) reactions, we will use shRNAs against factors in each pathway to establish whether POLQ inactivation is epistatic with either of them. In addition, the role of POLQ in bypassing telomeric G-quadruplexes during telomere replication will be investigated. Such a role could be reminiscent of the recently reported POLQ ability to overcome similar replication barriers in C. elegans. Together, these studies will shed light on how the recently-discovered POLQ-dependent pathway of DNA repair contributes to telomere integrity and tumour suppression. In the long term, this will enable us to develop systematic and comprehensive approaches of POLQ targeting for therapeutic purposes.