Haploinsufficiency of the Mus81-Eme1 endonuclease activates the intra-S-phase and G2/M checkpoints and promotes rereplication in human cells

Abstract

The Mus81–Eme1 complex is a structure-specific endonuclease that preferentially cleaves nicked Holliday junctions, 3′-flap structures and aberrant replication fork structures. Mus81−/− mice have been shown to exhibit spontaneous chromosomal aberrations and, in one of two models, a predisposition to cancers. The molecular mechanisms underlying its role in chromosome integrity, however, are largely unknown. To clarify the role of Mus81 in human cells, we deleted the gene in the human colon cancer cell line HCT116 by gene targeting. Here we demonstrate that Mus81 confers resistance to DNA crosslinking agents and slight resistance to other DNA-damaging agents. Mus81 deficiency spontaneously promotes chromosome damage such as breaks and activates the intra-S-phase checkpoint through the ATM-Chk1/Chk2 pathways. Furthermore, Mus81 deficiency activates the G2/M checkpoint through the ATM-Chk2 pathway and promotes DNA rereplication. Increased rereplication is reversed by the ectopic expression of Cdk1. Haploinsufficiency of Mus81 or Eme1 also causes similar phenotypes. These findings suggest that a complex network of the checkpoint pathways that respond to DNA double-strand breaks may participate in some of the phenotypes associated with Mus81 or Eme1 deficiency.

The EGFR-STAT3 Oncogenic Pathway Up-regulates the Eme1 Endonuclease to Reduce DNA Damage after Topoisomerase I Inhibition

“…As a consequence, the overexpression of the Mus81-Eme1 complex confers resistance to agents that block replication forks such as hydroxyurea and topoisomerase I inhibitors. In addition, its inactivation has been shown to induce genomic instability and enhance the sensitivity to DNA cross-linking agents (28,32). We therefore propose that a high expression of Eme1 due to STAT3 activation represents an intrinsic drug resistance program that allows EGFR-scr-expressing tumors to respond to topoisomerase I inhibitors.…”

Functional Interplay of p53 and Mus81 in DNA Damage Responses and Cancer

“…evidence describing the molecular mechanisms that underlie Mus81-mediated DNA repair is lacking (9,18,20). As such, we first addressed if Mus81 expression is altered in response to DNA damage by examining the effect of the ICL agent MMC on MEFs (Fig.…”

“…Mus81 deficiency increases sensitivity to DNA-damaging agents in yeast and mammals (8,9,(18)(19)(20). In particular, loss of mammalian Mus81 results in hypersensitivity to MMC.…”

“…Cellular phenotypes of Mus81-deficient murine and human cells include exquisite sensitivity to ICL agents and elevated levels of spontaneous and MMC-induced genomic instability (18)(19)(20). Importantly, our previous study showed that Mus81 +/À and Mus81 À/À mice spontaneously develop tumors, thus establishing Mus81 as a novel haploinsufficient tumor suppressor gene (18).…”

SLX4-SLX1 Protein-independent Down-regulation of MUS81-EME1 Protein by HIV-1 Viral Protein R (Vpr)

“…Purified recombinant MUS81-EME1 recognizes and preferentially cleaves several DNA structures, including 3Ј-flap, aberrant replication forks, and nicked Holliday junctions (45,48). Loss of MUS81 or EME1, or their disruption, results in genomic instability and hypersensitivity to DNA cross-linking agents such as mitomycin C and platinum compounds, due to DNA repair defects (49,50).…”