Alternative end-joining results in smaller deletions in heterochromatin relative to euchromatin

Abstract

Pericentromeric heterochromatin is highly enriched for repetitive sequences prone to aberrant recombination. Previous studies showed that homologous recombination (HR) repair is uniquely regulated in this domain to enable ‘safe’ repair while preventing aberrant recombination. InDrosophilacells, DNA double-strand breaks (DSBs) relocalize to the nuclear periphery through nuclear actin-driven directed motions before recruiting the strand invasion protein Rad51 and completing HR. End-joining (EJ) repair also occurs with high frequency in heterochromatin of fly tissues, but how different EJ pathways operate in heterochromatin remains uncharacterized. Here, we induce DSBs in single euchromatic and heterochromatic sites using the DR-whitereporter and I-SceI expression in spermatogonia. We detect higher frequency of HR repair in heterochromatic insertions, relative to euchromatin. Sequencing of repair outcomes reveals the use of distinct EJ pathways across different euchromatic and heterochromatic sites. Interestingly, synthesis-dependent michrohomology-mediated end joining (SD-MMEJ) appears differentially regulated in the two domains, with a preferential use of motifs close to the cut site in heterochromatin relative to euchromatin, resulting in smaller deletions. Together, these studies establish a new approach to study repair outcomes in fly tissues, and support the conclusion that heterochromatin uses more HR and less mutagenic EJ repair relative to euchromatin.

The impact of developmental stage, tissue type, and sex on DNA double-strand break repair in Drosophila melanogaster

“…Lastly, DSB repair may persist beyond the premeiotic germline (e.g., I-SceI transcripts that may persist after homologs segregate in meiosis I) which may impact repair outcomes. Studies limiting expression of I-SceI to the premeiotic germline may provide further insight to the effect of sex on repair in germline tissues [ 64 ]. Further studies are required to elucidate the role that Drosophila p53 (or other factors) may play in suppressing HR repair in females.…”