van Gent, D. C. | Search bioRxiv

bioRxiv2019

Low Dose Ionizing Radiation Strongly Stimulates Insertional Mutagenesis in a γH2AX Dependent Manner

Extrachromosomal DNA can integrate into the genome with no sequence specificity producing an insertional mutation. This process, which is referred to as random integration (RI), requires a double stranded break (DSB) in the genome. Inducing DSBs by various means, including ionizing radiation, increases the frequency of integration. Here we report that non-lethal physiologically relevant doses of ionizing radiation (10-100 mGy), within the range produced by medical imaging equipment, stimulate RI of transfected and viral episomal DNA in human and mouse cells with an extremely high efficiency. Genetic analysis of stimulated RI (S-RI) revealed that it is distinct from the background RI, requires histone H2AX S139 phosphorylation ({gamma}H2AX) and is not reduced by DNA polymerase {theta} (Polq) inactivation. S-RI efficiency was unaffected by the main DSB repair pathway (homologous recombination and non-homologous end joining) disruptions, but double deficiency in MDC1 and 53BP1 phenocopies {gamma}H2AX inactivation. The robust responsiveness of S-RI to physiological amounts of DSBs has implications for radiation risk assessment and can be exploited for extremely sensitive, macroscopic and direct detection of DSB-induced mutations.

molecular biology↗

bioRxiv2018

HSF2BP Negatively Regulates Homologous Recombination in DNA Interstrand Crosslink Repair in Human Cells by Direct Interaction With BRCA2

The tumor suppressor BRCA2 is essential for homologous recombination, replication fork stability and DNA interstrand crosslink (ICL) repair in vertebrates. We show that a functionally uncharacterized protein, HSF2BP, is involved in a novel, direct and highly evolutionarily conserved interaction with BRCA2. Although HSF2BP was previously described as testis-specific, we find it is expressed in mouse ES cells, in human cancer cell lines, and in tumor samples. Elevated levels of HSF2BP sensitize human cells to ICL-inducing agents (mitomycin C and cisplatin) and PARP inhibitors, resulting in a phenotype characteristic of cells from Fanconi anemia (FA) patients. We biochemically recapitulate the suppression of ICL repair and establish that excess HSF2BP specifically compromises homologous recombination by preventing BRCA2 and RAD51 loading at the ICL. As increased ectopic expression of HSF2BP occurs naturally, we suggest that it can be considered as a causative agent in FA and a source of cancer-promoting genomic instability.

molecular biology↗