Ce 4+ induced down-regulation of ERK-like MAPK and activation of nucleases during the apoptosis of cultured Taxus cuspidata cells

Abstract

Ce 4+ (Ce(NH 4) 2(NO 3) 6) at 1 mM induces apoptosis of suspension cultures of Taxus cuspidata cells; however, the underlying signal mechanisms are unknown. We show here that a 46-kDa ERK (extracellular signal-regulated kinase)-like MAPK appears to be down-regulated at 4 h, and remains at low levels for up to 48 h. An inhibitor of superoxide anions ( O 2 - ) generation, diphenyl iodonium (DPI) successfully blocks down-regulation of ERK-like MAPK and degradation of DNA. Moreover, a 41-kDa p38-like MAPK activity remains unchanged from 0.5 to 48 h. The p38 inhibitor SB202190 effectively inhibits p38-like MAPK activity, however, SB202190 fails to modify the apoptotic rate at concentrations up to 100 μM. Three nuclease (34-kDa, 22-kDa and 20-kDa) activities are profoundly enhanced in Ce 4+-induced T. cuspidata cells. They have an optimum pH at 6.8, and are stimulated by Ca 2+/Mg 2+. Caspase-3 inhibitor, Ac-DEVD-CHO, does not attenuate the 34-kDa nuclease activity, but inhibits the 22-kDa and the 20-kDa nuclease activities. In addition, inhibition of O 2 - generation by DPI significantly reduces the three nuclease activities. In conclusion, the present study suggests that down-regulation of ERK-like MAPK, burst of O 2 - , activation of caspase-3-like and induction of three nucleases as the key signaling events mediating apoptosis in Ce 4+-induced cultured T. cuspidata cells.