Mutational signatures of non‐homologous and polymerase theta‐mediated end‐joining in embryonic stem cells. (27th October 2017)

• Record Type: Journal Article
• Authors: Schimmel, Joost; Kool, Hanneke; van Schendel, Robin; Tijsterman, Marcel
• Is Part Of: EMBO journal. Volume 36:Number 24(2017)
• Page Start: 3634
• Page End: 3649
• Publication Date: 2017-10-27
• Subjects: cNHEJ -- CRISPR/Cas9 -- double‐strand break repair -- embryonic stem cells -- polymerase theta‐mediated end joining; Molecular biology -- Periodicals; 572.805
• DOI: 10.15252/embj.201796948
• Languages: English
• ISSNs: 0261-4189

Abstract

Abstract: Cells employ potentially mutagenic DNA repair mechanisms to avoid the detrimental effects of chromosome breaks on cell survival. While classical non‐homologous end‐joining (cNHEJ) is largely error‐free, alternative end‐joining pathways have been described that are intrinsically mutagenic. Which end‐joining mechanisms operate in germ and embryonic cells and thus contribute to heritable mutations found in congenital diseases is, however, still largely elusive. Here, we determined the genetic requirements for the repair of CRISPR/Cas9‐induced chromosomal breaks of different configurations, and establish the mutational consequences. We find that cNHEJ and polymerase theta‐mediated end‐joining (TMEJ) act both parallel and redundant in mouse embryonic stem cells and account for virtually all end‐joining activity. Surprisingly, mutagenic repair by polymerase theta (Pol θ, encoded by the Polq gene) is most prevalent for blunt double‐strand breaks (DSBs), while cNHEJ dictates mutagenic repair of DSBs with protruding ends, in which the cNHEJ polymerases lambda and mu play minor roles. We conclude that cNHEJ‐dependent repair of DSBs with protruding ends can explain de novo formation of tandem duplications in mammalian genomes. Synopsis: Embryonic stem cells utilize two distinct mutagenic end‐joining mechanisms to repair chromosomal breaks. The configuration of the break dictates repair‐pathway choice, resulting in different repair outcomes, which can explain mutational signatures observed in human disease and evolution. Theta‐mediated end‐joining (TMEJ) acts as a first line defense mechanism for chromosomal breaks in mouse embryonic stem cells. Mutational signatures obtained in various human disease can be explained by TMEJ‐dependent repair of chromosomal breaks. Classical non‐homologous end‐joining (cNHEJ) of chromosomal breaks with protruding ends can explain the formation of tandem duplications in mammalian genomes. The introduction of one single chromosomal break induces lethality in cells that lack both TMEJ and cNHEJ. Abstract : DNA double‐strand break configuration dictates the choice between two distinct error‐prone repair pathways in mouse embryonic stem cells, resulting in different repair outcomes.