title: Transcript-RNA-templated DNA recombination and repair.
abstract: Homologous recombination is a molecular process that has multiple important roles in DNA metabolism, both for DNA repair and genetic variation in all forms of life. Generally, homologous recombination involves the exchange of genetic information between two identical or nearly identical DNA molecules; however, homologous recombination can also occur between RNA molecules, as shown for RNA viruses. Previous research showed that synthetic RNA oligonucleotides can act as templates for DNA double-strand break (DSB) repair in yeast and human cells, and artificial long RNA templates injected in ciliate cells can guide genomic rearrangements. Here we report that endogenous transcript RNA mediates homologous recombination with chromosomal DNA in yeast Saccharomyces cerevisiae. We developed a system to detect the events of homologous recombination initiated by transcript RNA following the repair of a chromosomal DSB occurring either in a homologous but remote locus, or in the same transcript-generating locus in reverse-transcription-defective yeast strains. We found that RNA-DNA recombination is blocked by ribonucleases H1 and H2. In the presence of H-type ribonucleases, DSB repair proceeds through a complementary DNA intermediate, whereas in their absence, it proceeds directly through RNA. The proximity of the transcript to its chromosomal DNA partner in the same locus facilitates Rad52-driven homologous recombination during DSB repair. We demonstrate that yeast and human Rad52 proteins efficiently catalyse annealing of RNA to a DSB-like DNA end in vitro. Our results reveal a novel mechanism of homologous recombination and DNA repair in which transcript RNA is used as a template for DSB repair. Thus, considering the abundance of RNA transcripts in cells, RNA may have a marked impact on genomic stability and plasticity.
authors.affiliation: School of Biology, Georgia Institute of Technology, Atlanta, Georgia 30332, USA.
authors.forename: Havva
authors.initials: H
authors.lastname: Keskin
authors.affiliation: 1] School of Biology, Georgia Institute of Technology, Atlanta, Georgia 30332, USA [2] Division of Computational Biomedicine, Boston University School of Medicine, Boston, Massachusetts 02118, USA.
authors.forename: Ying
authors.initials: Y
authors.lastname: Shen
authors.affiliation: Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, Pennsylvania 19102, USA.
authors.forename: Fei
authors.initials: F
authors.lastname: Huang
authors.affiliation: Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, Pennsylvania 19102, USA.
authors.forename: Mikir
authors.initials: M
authors.lastname: Patel
authors.affiliation: School of Biology, Georgia Institute of Technology, Atlanta, Georgia 30332, USA.
authors.forename: Taehwan
authors.initials: T
authors.lastname: Yang
authors.affiliation: School of Biology, Georgia Institute of Technology, Atlanta, Georgia 30332, USA.
authors.forename: Katie
authors.initials: K
authors.lastname: Ashley
authors.affiliation: Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, Pennsylvania 19102, USA.
authors.forename: Alexander V
authors.initials: AV
authors.lastname: Mazin
authors.affiliation: School of Biology, Georgia Institute of Technology, Atlanta, Georgia 30332, USA.
authors.forename: Francesca
authors.initials: F
authors.lastname: Storici
mesh_terms: D015825:Chromosomes, Fungal / Q000235:genetics; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair / Q000235:genetics*; D042822:Genomic Instability / Q000235:genetics; D059765:Homologous Recombination / Q000235:genetics*; D006801:Humans; D008957:Models, Genetic; D012313:RNA / Q000235:genetics*; D051721:Rad52 DNA Repair and Recombination Protein / Q000378:metabolism; D016914:Ribonuclease H / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D013698:Templates, Genetic; D014158:Transcription, Genetic / Q000235:genetics*
chemical_list: C497197:RAD52 protein, S cerevisiae; C497194:RAD52 protein, human; D051721:Rad52 DNA Repair and Recombination Protein; D029701:Saccharomyces cerevisiae Proteins; D012313:RNA; C091139:ribonuclease HII; D016914:Ribonuclease H; C078856:ribonuclease HI