Double-strand DNA breaks are mainly repaired by the homologous recombination pathway in early developing swine embryos

Abstract

DNA double-strand breaks (DSBs) are less frequent than single-strand breaks but have more harmful consequences on cell survival and physiology. Homologous recombination (HR) and nonhomologous end-joining (NHEJ) are thetwomainpathways that are responsible forDSBrepair ineukaryotic cells, but their importance for the preservation of genome stability in totipotent blastomeres of early developing embryos has not been determined. In this study, we observed that the chemical inhibition of HR or both pathways, but not NHEJ alone, increased the number of DSBs, reduced embryo development to the blastocyst stage, and resulted in embryos with higher proportions of apoptotic cells. Targeted knockdown of ATM (ataxia telangiectasia mutated) and ATR (ataxia telangiectasia andRad3 related;HRregulators) andDNA-dependent proteinkinase (NHEJregulator)mRNAsrevealed that the attenuation of HR or both HR and NHEJ regulators severely impaired blastocyst formation and quality. Attenuation of ATM alone resulted in a higher incidence of DSBs, lower development and embryo quality, and increasedmRNA abundance of genes that are involved in either repair pathway. These findings indicate that HR is themain pathway responsible for the promotion ofDSB repair in early developing embryos, and thatATMseems to be more important thanATRin the regulationof theHRpathway inmammalianembryos.