Hydrogen Peroxide Stimulates c-Src-mediated Big Mitogen-activated Protein Kinase 1 (BMK1) and the MEF2C Signaling Pathway in PC12 Cells

Reactive oxygen species, generated by reduction-oxidation (redox) reactions, have been recognized as one of the major mediators of ischemia and reperfusion injury in the brain. Reactive oxygen species-induced cerebral events are attributable, in part, to the change in intracellular signaling molecules including mitogen-activated protein (MAP) kinases. Big MAP kinase 1 (BMK1), also known as ERK5, is a newly identified member of the MAP kinase family and has been reported to be sensitive to oxidative stress. In the present study, we examined the effect of H₂O₂ on BMK1 activity in PC12 cells, and we investigated the pathophysiological implication of BMK1. Findings showed that BMK1 was rapidly and significantly activated by H₂O₂ in a concentration-dependent manner in PC12 cells. BMK1 activation by H₂O₂was inhibited by both PD98059 and U0126, which were reported to inhibit MEK5 as well as MEK1/2. c-Src was suggested to be involved in BMK1 activation from the experiments with herbimycin A and PP2, specific inhibitors of Src family kinases. Transfection of kinase-inactive Src also inhibited H₂O₂-induced BMK1 activation. In addition, H₂O₂ treatment of cells induced an enhancement of DNA binding activity of MEF2C, a downstream transcription factor of BMK1 in PC12 cells. Finally, pretreatment of cells with PD98059 and U0126 resulted in an increase in cell death including apoptosis by H₂O₂ in ERK1/2 down-regulated cells as well as in intact PC12 cells. These findings suggest that c-Src mediated BMK1 activation by H₂O₂ may counteract ischemic cellular damage probably through the activation of MEF2C transcription factor.