Mammalian Rad9 Plays a Role in Telomere Stability, S- and G₂-Phase-Specific Cell Survival, and Homologous Recombinational Repair

Abstract

The protein products of several rad checkpoint genes of Schizosaccharomyces pombe (rad1؉ , rad26 ؉ , and hus1 ؉ ) play crucial roles in sensing changes in DNA structure, and several function in the maintenance of telomeres. When the mammalian homologue of S. pombe Rad9 was inactivated, increases in chromosome end-to-end associations and frequency of telomere loss were observed. This telomere instability correlated with enhanced S-and G 2 -phase-specific cell killing, delayed kinetics of ␥-H2AX focus appearance and disappearance, and reduced chromosomal repair after ionizing radiation (IR) exposure, suggesting that Rad9 plays a role in cell cycle phase-specific DNA damage repair. Furthermore, mammalian Rad9 interacted with Rad51, and inactivation of mammalian Rad9 also resulted in decreased homologous recombinational (HR) repair, which occurs predominantly in the S and G 2 phases of the cell cycle. Together, these findings provide evidence of roles for mammalian Rad9 in telomere stability and HR repair as a mechanism for promoting cell survival after IR exposure.

Human Exonuclease 5 Is a Novel Sliding Exonuclease Required for Genome Stability

“…Human Cell Growth and Transfection-Human 293 and GM5849 cells were maintained by previously published procedures (13). Full-length FLAG-tagged hEXO5 cDNA cloned in pcDNA3.1 was transfected into cells.…”

Clamping down on mammalian meiosis

“…For example, siRNA knockdown of Hus1 expression was shown to decrease the efficiency of homologous recombination in response to ionizing radiation (IR)-induced DNA damage, and HUS1-deficient mouse cells are hypersensitive to IR despite intact nonhomologous end joining (NHEJ). 47 Mammalian RAD9A interacts with the RAD51 recombinase, 3 and Rad9a inactivation leads to decreased repair of DSBs by homologous recombination and inefficient exit from G 2 phase following IR exposure. 3,48 Similarly, RAD9A-deficient mouse embryonic stem cells initiate but do not maintain IR-induced G 2 delay.…”

“…47 Mammalian RAD9A interacts with the RAD51 recombinase, 3 and Rad9a inactivation leads to decreased repair of DSBs by homologous recombination and inefficient exit from G 2 phase following IR exposure. 3,48 Similarly, RAD9A-deficient mouse embryonic stem cells initiate but do not maintain IR-induced G 2 delay. 44 Immunoglobulin gene conversion in Rad9a −/− chicken DT40 cells is substantially reduced in a manner similar to Rad51 or Brca2 mutants with known defects in homologous recombination.…”

“…Roles for 9-1-1 in DNA repair are less well understood; however, physical and functional interactions with many DNA repair proteins have been reported, including a variety of DNA polymerases, glycosylases, nucleases, and the RAD51 recombinase. [1][2][3] These interactions implicate 9-1-1 function in DNA repair pathways as diverse as base excision repair, nucleotide excision repair, mismatch repair, and homologous recombination, most likely in recognizing DNA damage and coordinating recruitment of critical repair factors.…”

Ubiquitin-specific Peptidase 20 Regulates Rad17 Stability, Checkpoint Kinase 1 Phosphorylation and DNA Repair by Homologous Recombination

“…Each of Rad17 (7,16), the 9-1-1 clamp (17), Chk1 (18), and PLK1 (19) has been reported to be involved in the HR repair of DSBs. The 9-1-1 clamp interacts with Rad51 (17), and Chk1 phosphorylates this key HR protein (18). HR proceeds when the DSB end is processed by exonucleases to generate ssDNA that binds Rad51 protein and forms a nucleofilament, which invades the homologous sequence to repair the damage (15).…”