Loss of DNA mismatch repair imparts defective cdc2 signaling and G2 arrest responses without altering survival after ionizing radiation

T. Yan, J. E. Schupp, H. S. Hwang, M. W. Wagner, S. E. Berry, S. Strickfaden, M. L. Veigl, W. D. Sedwick, D. A. Boothman, T. J. Kinsella

Research output: Contribution to journalArticle

67 Scopus citations

Abstract

Our previous data demonstrated that cells deficient in MutL homologue-1 (MLH1) expression had a reduced and shorter G2 arrest after high-dose-rate ionizing radiation (IR), suggesting that the mismatch repair (MMR) system mediates this cell cycle checkpoint. We confirmed this observation using two additional isogenetically matched human MLH1 (hMLH1)-deficient and -proficient human tumor cell systems: human ovarian cancer cells, A2780/CP70, with or without ectopically expressed hMLH1, and human colorectal carcinoma cells, RKO, with or without azacytidine treatment to reexpress hMLH1. We also examined matched MutS homologue-2 (hMSH2)-deficient and -proficient human endometrial carcinoma HEC59 cell lines to determine whether hMSH2, and MMR in general, is involved in IR-related G2 arrest responses. As in MLH1-deficient cells, cells lacking hMSH2 demonstrated a similarly altered G2 arrest in response to IR (6 Gy). These differences in IR-induced G2 arrest between MMR-proficient and -deficient cells were found regardless of whether synchronized cells were irradiated in G0/G1 or S phase, indicating that MMR indeed dramatically affects the G2-M checkpoint arrest. However, unlike the MMR-dependent damage tolerance response to 6-thioguanine exposures, no significant difference in the clonogenic survival of MMR-deficient cells compared with MMR-proficient cells was noted after high-dose-rate IR. In an attempt to define the signal transduction mechanisms responsible for MMR-mediated G2 arrest, we examined the levels of tyrosine 15 phosphorylation of cdc2 (phospho-Tyr15 -cdc2), a key regulator of the G2-M transition. Increased phospho-Tyr15-cdc2 levels were observed in both MMR-proficient and -deficient cell lines after IR. However, the levels of the phospho-Tyr15-cdc2 rapidly decreased in MMR (hMLH1 or hMSH2)-deficient cell lines at times coincident with progress from the IR-induced G2 arrest through M phase. Thus, differences in the levels of phospho-Tyr15-cdc2 after high-dose-rate IR correspond temporally with the observed differences in the IR-induced G2 arrest, suggesting that MMR proteins may exert their effect on IR-induced G2 arrest by signaling the cdc2 pathway. Although MMR status does not significantly affect the survival of cells after high-dose-rate IR, it seems to regulate the G2-M checkpoint and might affect overall mutation rates.

Original languageEnglish (US)
Pages (from-to)8290-8297
Number of pages8
JournalCancer Research
Volume61
Issue number22
Publication statusPublished - Nov 15 2001

    Fingerprint

ASJC Scopus subject areas

  • Cancer Research
  • Oncology

Cite this

Yan, T., Schupp, J. E., Hwang, H. S., Wagner, M. W., Berry, S. E., Strickfaden, S., ... Kinsella, T. J. (2001). Loss of DNA mismatch repair imparts defective cdc2 signaling and G2 arrest responses without altering survival after ionizing radiation. Cancer Research, 61(22), 8290-8297.