BLM has early and late functions in homologous recombination repair in mouse embryonic stem cells

TY - JOUR

T1 - BLM has early and late functions in homologous recombination repair in mouse embryonic stem cells

AU - Chu, W K

AU - Hanada, K

AU - Kanaar, R

AU - Hickson, I D

PY - 2010/8/19

Y1 - 2010/8/19

N2 - BLM is a RecQ family helicase that is defective in individuals with the cancer predisposition disorder, Bloom's syndrome (BS). At the cellular level, BS is characterized by hyper-recombination manifested as excessive sister chromatid exchange and loss of heterozygosity. However, the precise function of BLM remains unclear. Multiple roles have been proposed for BLM in the homologous recombination (HR) repair pathway, including 'early' functions, such as the stimulation of resection of DNA double-strand break ends or displacement of the invading strand of DNA displacement loops, and 'late' roles, such as dissolution of double Holliday junctions. However, most of the evidence for these putative roles comes from in vitro biochemical data. In this study, we report the characterization of mouse embryonic stem cells with disruption of Blm and/or Rad54 genes. We show that Blm has roles both upstream and downstream of the Rad54 protein, a core HR factor. Disruption of Rad54 in the Blm-mutant background reduced the elevated level of gene targeting and of sister chromatid exchanges, implying that Blm primarily functions downstream of Rad54 in the HR pathway. Conversely, however, mutation of Blm in Rad54(-/-) cells rescued their mitomycin C (MMC) sensitivity, and decreased both the level of DNA damage and cell cycle perturbation induced by MMC, suggesting an early role for Blm. Our data are consistent with Blm having at least two roles in HR repair in mammalian cells.

AB - BLM is a RecQ family helicase that is defective in individuals with the cancer predisposition disorder, Bloom's syndrome (BS). At the cellular level, BS is characterized by hyper-recombination manifested as excessive sister chromatid exchange and loss of heterozygosity. However, the precise function of BLM remains unclear. Multiple roles have been proposed for BLM in the homologous recombination (HR) repair pathway, including 'early' functions, such as the stimulation of resection of DNA double-strand break ends or displacement of the invading strand of DNA displacement loops, and 'late' roles, such as dissolution of double Holliday junctions. However, most of the evidence for these putative roles comes from in vitro biochemical data. In this study, we report the characterization of mouse embryonic stem cells with disruption of Blm and/or Rad54 genes. We show that Blm has roles both upstream and downstream of the Rad54 protein, a core HR factor. Disruption of Rad54 in the Blm-mutant background reduced the elevated level of gene targeting and of sister chromatid exchanges, implying that Blm primarily functions downstream of Rad54 in the HR pathway. Conversely, however, mutation of Blm in Rad54(-/-) cells rescued their mitomycin C (MMC) sensitivity, and decreased both the level of DNA damage and cell cycle perturbation induced by MMC, suggesting an early role for Blm. Our data are consistent with Blm having at least two roles in HR repair in mammalian cells.

KW - Animals

KW - Cell Line, Tumor

KW - DNA Damage

KW - DNA Helicases

KW - DNA Repair

KW - DNA Replication

KW - DNA-Binding Proteins

KW - Embryonic Stem Cells

KW - Gene Targeting

KW - Immunoblotting

KW - Mice

KW - Nuclear Proteins

KW - RecQ Helicases

KW - Recombinant Proteins

KW - Recombination, Genetic

KW - Sister Chromatid Exchange

U2 - 10.1038/onc.2010.214

DO - 10.1038/onc.2010.214

M3 - Journal article

C2 - 20531307

SN - 0950-9232

VL - 29

SP - 4705

EP - 4714

JO - Oncogene

JF - Oncogene

IS - 33

ER -