Project description:Data from Excision-seq experiments to map deoxyuridine and pyrimidine dimers in S. cerevisiae and E. coli For uracil mapping in pre-digestion Excision-seq, uracil was excised from genomic DNA from dut ung yeast and bacteria yielding double-stranded DNA fragments. Adapters were ligated to these fragments for Illumina library preparation. Using this method, the number of reads at a genomic location corresponds to the quantity of dU at that location. In post-digestion Excision-seq, uracil-containing library fragments are destroyed by UDG treatment, thus coverage is inversely proportional to uracil content. For pyrimidine dimer Excision-seq, yeast were irradiated with high doses of UVC light to generate a large number of DNA modifications. Using the excision repair enzyme UVDE from S. pombe, we excised pyrimidine dimers, cutting the phosphodieseter bond to release small double stranded fragments. These fragments were repaired with either CPD photolyase or 6-4 photolyase to generate independent libraries for each modification type. Illumina adapters were ligated to these fragments for library preparation. Using this method the number of reads at a genomic location corresponds to the quantity of the 3' pyrimidine of the dimer.
Project description:Uncovering novel mechanisms controlling DNA damage response (DDR) is essential to better understand the molecular features of high-grade serous ovarian cancers (HGSOC), which remain a clinical challenge. Here we demonstrate that the glutaminyl-tRNA-synthetase QARS is a new key player in DDR. We show that QARS is regulated at translational level by the MAPK pathway in HGSOC and controls DDR through a translation-independent mechanism. Analyses combining RNA sequencing, immunohistochemistry, proteomic data and functional assays from HGSOC patient cohorts and relevant cellular models reveal that QARS promotes homologous recombination (HR)-mediated DNA repair in HGSOC, a function that requires its enzymatic activity. Interestingly, we uncover that QARS is located in the nucleus. Moreover, DNA damage increases QARS protein levels at the chromatin and its interaction with nuclear autoantigen SP100 and Dioxyuridine Triphosphatase DUT, involved in DNA break sensing and DNA integrity. Our data thus highlight a novel nuclear function and mechanism by which QARS enhances DDR in HGSOC.