Genome-wide Analysis of DNA-PK-bound MRN Cleavage Products Supports a Sequential Model of DSB Repair Pathway Choice

Overview

Journal Nat Commun

Specialty Biology

Date 2023 Sep 16

PMID 37717054

Authors

Rajashree A Deshpande

Alberto Marin-Gonzalez

Hannah K Barnes

Phillip R Woolley

Taekjip Ha

Tanya T Paull

Affiliations

Soon will be listed here.

Abstract

The Mre11-Rad50-Nbs1 (MRN) complex recognizes and processes DNA double-strand breaks for homologous recombination by performing short-range removal of 5' strands. Endonucleolytic processing by MRN requires a stably bound protein at the break site-a role we postulate is played by DNA-dependent protein kinase (DNA-PK) in mammals. Here we interrogate sites of MRN-dependent processing by identifying sites of CtIP association and by sequencing DNA-PK-bound DNA fragments that are products of MRN cleavage. These intermediates are generated most efficiently when DNA-PK is catalytically blocked, yielding products within 200 bp of the break site, whereas DNA-PK products in the absence of kinase inhibition show greater dispersal. Use of light-activated Cas9 to induce breaks facilitates temporal resolution of DNA-PK and Mre11 binding, showing that both complexes bind to DNA ends before release of DNA-PK-bound products. These results support a sequential model of double-strand break repair involving collaborative interactions between homologous and non-homologous repair complexes.

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