Haploinsufficiency of the Mus81-Eme1 endonuclease activates the intra-S-phase and G2/M checkpoints and promotes rereplication in human cells.
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.
Hiyama
Takashi
T
Department of Human Genetics, Hiroshima University 1-2-3 Kasumi, Minami-ku, Hiroshima 734-8553, Japan.
Katsura
Mari
M
Yoshihara
Takashi
T
Ishida
Mari
M
Kinomura
Aiko
A
Tonda
Tetsuji
T
Asahara
Toshimasa
T
Miyagawa
Kiyoshi
K
D016203:CDC2 Protein Kinase / Q000378:metabolism; D002453:Cell Cycle; D002460:Cell Line; D043171:Chromosomal Instability; D004249:DNA Damage; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology*; D004706:Endodeoxyribonucleases / Q000235:genetics / Q000502:physiology*; D004720:Endonucleases / Q000235:genetics / Q000502:physiology*; D016195:G2 Phase; D017353:Gene Deletion; D018390:Gene Targeting; D006579:Heterozygote; D006801:Humans; D008938:Mitosis; D011123:Polyploidy; D016196:S Phase
D004268:DNA-Binding Proteins; D016203:CDC2 Protein Kinase; C507947:Eme1 protein, human; D004706:Endodeoxyribonucleases; D004720:Endonucleases; C498772:MUS81 protein, human