MAPK dependence of DNA damage repair: ionizing radiation and the induction of expression of the DNA repair genes XRCC1 and ERCC1 in DU145 human prostate carcinoma cells in a MEK1/2 dependent fashion

Purpose: To examine the role of mitogen-activated protein kinase (MAPK) signalling on the induction by ionizing radiation of the nucleotide excision repair gene (ERCC1), the X-ray cross-complementing group 1 protein (XRCC1) and the repair of radiation-induced DNA damage. Materials and methods: The expression of ERCC1 and XRCC1 was examined in DU145 human prostate cancer cells following exposure to ionizing radiation. We characterized the MAPK dependence of this expression through RT-PCR analysis, Western analysis, transcription inhibition and measurement of the activation of each promoter. Pre-exposure with the specific MEK1/2 inhibitor PD980059 (10 muM) was used to blunt radiation induction of MAPK without suppressing basal levels of MAPK activity. In addition, we examined the MA-PK dependence of DNA damage repair by measuring radiation-induced micronucleus formation and the removal of and nicking activity associated with AP sites. Results: Irradiation caused a time-dependent, MAPK-dependent increase in the protein levels of both ERCC1 and XRCC1. For each gene product, the protein level increase followed an increase in mRNA, which also was MAPK-dependent. Radiation also enhanced the activities of the ERCC1 and XRCC1 promoters in an MAPK-dependent fashion. Inhibition of transcription by DRB abolished the radiation-induced increase of ERCC1 and XRCC1 proteins. Inhibition of radiation-induced MAPK also diminished the ability of DU145 cells to remove AP sites and increased the number of cells displaying micronuclei following radiation exposure. Conclusions: These findings demonstrate a role for radiation-induced MAPK signalling in the regulation of DNA repair enzyme levels and DNA repair. Radiation-induced protein expression of ERCC1 and XRCC1 appears to require de novo transcription. These data suggest a significant role for MAPK signalling in the early response to DNA damage caused by ionizing radiation.