title: Multiple mechanisms contribute to double-strand break repair at rereplication forks in Drosophila follicle cells.
abstract: Rereplication generates double-strand breaks (DSBs) at sites of fork collisions and causes genomic damage, including repeat instability and chromosomal aberrations. However, the primary mechanism used to repair rereplication DSBs varies across different experimental systems. In Drosophila follicle cells, developmentally regulated rereplication is used to amplify six genomic regions, two of which contain genes encoding eggshell proteins. We have exploited this system to test the roles of several DSB repair pathways during rereplication, using fork progression as a readout for DSB repair efficiency. Here we show that a null mutation in the microhomology-mediated end-joining (MMEJ) component, polymerase θ/mutagen-sensitive 308 (mus308), exhibits a sporadic thin eggshell phenotype and reduced chorion gene expression. Unlike other thin eggshell mutants, mus308 displays normal origin firing but reduced fork progression at two regions of rereplication. We also find that MMEJ compensates for loss of nonhomologous end joining to repair rereplication DSBs in a site-specific manner. Conversely, we show that fork progression is enhanced in the absence of both Drosophila Rad51 homologs, spindle-A and spindle-B, revealing homologous recombination is active and actually impairs fork movement during follicle cell rereplication. These results demonstrate that several DSB repair pathways are used during rereplication in the follicle cells and their contribution to productive fork progression is influenced by genomic position and repair pathway competition. Furthermore, our findings illustrate that specific rereplication DSB repair pathways can have major effects on cellular physiology, dependent upon genomic context.
authors.affiliation: Whitehead Institute for Biomedical Research, Cambridge, MA 02142.|Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02142.
authors.forename: Jessica L
authors.initials: JL
authors.lastname: Alexander
authors.affiliation: Department of Biology, Tufts University, Medford, MA 02155.
authors.forename: Kelly
authors.initials: K
authors.lastname: Beagan
authors.affiliation: Whitehead Institute for Biomedical Research, Cambridge, MA 02142; mitch.mcvey@tufts.edu weaver@wi.mit.edu.|Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02142.
authors.forename: Terry L
authors.initials: TL
authors.lastname: Orr-Weaver
authors.affiliation: Department of Biology, Tufts University, Medford, MA 02155 mitch.mcvey@tufts.edu weaver@wi.mit.edu.
authors.forename: Mitch
authors.initials: M
authors.lastname: McVey
mesh_terms: D000818:Animals; D053903:DNA Breaks, Double-Stranded; D059766:DNA End-Joining Repair / Q000235:genetics; D004261:DNA Replication / Q000235:genetics*; D029721:Drosophila Proteins / Q000235:genetics*; D004331:Drosophila melanogaster / Q000235:genetics / Q000254:growth & development; D004527:Egg Proteins / Q000235:genetics*; D005260:Female; D018507:Gene Expression Regulation, Developmental; D049750:Genome, Insect / Q000235:genetics; D059765:Homologous Recombination / Q000235:genetics*; D006080:Ovarian Follicle / Q000254:growth & development; D051135:Rad51 Recombinase / Q000235:genetics*; D015398:Signal Transduction / Q000235:genetics
chemical_list: D029721:Drosophila Proteins; D004527:Egg Proteins; C015753:chorion proteins; C114493:spn-B protein, Drosophila; D051135:Rad51 Recombinase; C495149:spn-A protein, Drosophila
keywords: DNA repair; NHEJ; break-induced replication; fork collision; oogenesis