Actin-Related Protein 6 (Arp6) Influences Double-Strand Break Repair in Yeast

Mohsen Hooshyar,Daniel Burnside,Matthew Jessulat,Houman Moteshareie,Adamo Young,Maryam Hajikarimlou,Andrew Schoenrock,Eshan Silva,Mohan Babu,Bahram Samanfar,Pedro Matilha Cherubini,Frank Dehne,Katayoun Omidi,Alexander Jesso,Ashkan Golshani,Jean-Simon Diallo,Megan Vanstone,Taylor Potter,Urvi Bhojoo

doi:10.3390/applmicrobiol1020017

Abstract

DNA double-strand breaks (DSBs) are the most deleterious form of DNA damage and are repaired through non-homologous end-joining (NHEJ) or homologous recombination (HR). Repair initiation, regulation and communication with signaling pathways require several histone-modifying and chromatin-remodeling complexes. In budding yeast, this involves three primary complexes: INO80-C, which is primarily associated with HR, SWR1-C, which promotes NHEJ, and RSC-C, which is involved in both pathways as well as the general DNA damage response. Here we identify ARP6 as a factor involved in DSB repair through an RSC-C-related pathway. The loss of ARP6 significantly reduces the NHEJ repair efficiency of linearized plasmids with cohesive ends, impairs the repair of chromosomal breaks, and sensitizes cells to DNA-damaging agents. Genetic interaction analysis indicates that ARP6, MRE11 and RSC-C function within the same pathway, and the overexpression of ARP6 rescues rsc2∆ and mre11∆ sensitivity to DNA-damaging agents. Double mutants of ARP6, and members of the INO80 and SWR1 complexes, cause a significant reduction in repair efficiency, suggesting that ARP6 functions independently of SWR1-C and INO80-C. These findings support a novel role for ARP6 in DSB repair that is independent of the SWR1 chromatin remodeling complex, through an apparent RSC-C and MRE11-associated DNA repair pathway.

Highlights

DNA double-strand breaks (DSBs) occur when both strands of the phosphodiester backbone are severed, and the integrity of the DNA molecule is compromised
Because Arp6 has been loosely linked to non-homologous end-joining (NHEJ), we first investigated if ARP6 influences the NHEJ pathway, using a series of DNA repair assays
When ARP6 mutants were presented with breaks containing 50 cohesive overhangs (XbaI digestion), repair efficiency was reduced to 41% compared to WT

Summary

Introduction

DNA double-strand breaks (DSBs) occur when both strands of the phosphodiester backbone are severed, and the integrity of the DNA molecule is compromised. Such breaks are regarded as the most genotoxic form of DNA damage, and can lead to genomic. Much research has focused on achieving a comprehensive understanding of DSB repair pathways because of their complex nature, biological importance and links to human diseases such as cancer [2,3]. Eukaryotic DSB repair proceeds primarily through one of two distinct pathways: homologous recombination (HR), which uses a homologous template and is considered error-free; or non-homologous end-joining (NHEJ), which directly re-joins the broken ends through an error-prone process

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