DNA-PK promotes DNA end resection at DNA double strand breaks in G0 cells

Abstract

DNA double-strand break (DSB) repair by homologous recombination is confined to the S and G2 phases of the cell cycle partly due to 53BP1 antagonizing DNA end resection in G1 phase and non-cycling quiescent (G0) cells where DSBs are predominately repaired by non-homologous end joining (NHEJ). Unexpectedly, we uncovered extensive MRE11- and CtIP-dependent DNA end resection at DSBs in G0 murine and human cells. A whole genome CRISPR/Cas9 screen revealed the DNA-dependent kinase (DNA-PK) complex as a key factor in promoting DNA end resection in G0 cells. In agreement, depletion of FBXL12, which promotes ubiquitylation and removal of the KU70/KU80 subunits of DNA-PK from DSBs, promotes even more extensive resection in G0 cells. In contrast, a requirement for DNA-PK in promoting DNA end resection in proliferating cells at the G1 or G2 phase of the cell cycle was not observed. Our findings establish that DNA-PK uniquely promotes DNA end resection in G0, but not in G1 or G2 phase cells, which has important implications for DNA DSB repair in quiescent cells.

DNA-PKcs Suppresses Illegitimate Chromosome Rearrangements

“…Though suppressing resection aligns with excluding A-EJ, promoting resection was surprising and through a mechanism that is not fully understood. 66 It is possible that the immature DNA-PK synapsis at DNA ends 64 provides a favorable platform for recruiting DNA polymerases, nucleases, and other factors to support an Artemis & resection-dependent NHEJ mechanism that was previously described in G1, but not G2, phase. 72,73 In this regard, ATM kinase can activate Artemis in the presence of an inhibited DNA-PKcs 39,74 and could be used to re-engage NHEJ-mediated ligation in the presence of Lig4.…”

“…Notably for JκCE baits, we observed a decreased range of resected joints with Lig4 -/- DNA-PKcs -/- #1/2 clones compared to Lig4 -/- alone, which aligns with recent studies demonstrating that DNA-PKcs promotes DSB resection. 66,67 In JκSE baits, Lig4 -/- DNA-PKcs -/- clones displayed varied effects on resection (Figure S13C). DNA-PKcs and Ku70 deletion also decreased MMEJ (>2bp) and correspondingly increased direct repair for all baits compared Lig4 -/- control (Figures S14A-S14D and S13D-S13G).…”

Meiotic DNA break resection and recombination rely on chromatin remodeler Fun30

“…Our findings here implicate Fun30 in this role in yeast. Chromatin remodeling contributes to resection in vegetative yeast and mammalian cells as well (Peritore et al 2021; Fowler et al 2022). However, unlike in vegetative cells, where Fun30 acts redundantly with other remodelers to promote resection specifically over relatively long distances (Chen et al 2012; Costelloe et al 2012; Eapen et al 2012), during meiosis Fun30 by itself strongly affects both initial (MRX/Sae2) and secondary (Exo1) resection steps.…”

Dynamic features of human mitochondrial DNA maintenance and transcription

“…In the nuclear genome, lesions are repaired by different DNA repair pathways depending on the nature of the lesion, phases of the cell cycle, and posttranslational modifications of the repair proteins ( Akbari et al, 2009 ; Mjelle et al, 2015 ; Carter and Parsons, 2016 ; Fowler et al, 2022 ). The main pathways include mismatch repair (MMR), nucleotide excision repair (NER), base excision repair (BER) and the double strand break (DSB) repair (DSBR) pathways homologous recombination (HR) or non-homologous end joining (NHEJ).…”