Activation of Rac1 increases c-Jun NH2-terminal kinase activity and DNA fragmentation in a calcium-dependent manner in rat myoblast cell line H9c2

We examined the role of intracellular Ca2+ in c-Jun NH2-terminal kinase (JN K) activation and DNA fragmentation in the rat myoblast cell line H9c2 usin g small GTP-binding protein Rad. A constitutively active mutant of Rad (V12 -Rac1) increased JNK-responsive gene expression 6-fold, although this incre ase was attenuated by the intracellular Ca2+ chelator BAPTA-AM. V12-Rac1 al so increased the number of DNA fragmentated cells. However, V12-Rac1-mediat ed JNK activation was not affected by BAPTA-AM as determined by direct meas urement of active forms, and V12-Rac1 did not affect intracellular Ca2+ con centration. These results suggest that Rad can activate JNK and induces cel l injury, but [Ca2+](i) is necessary for V12-Rac1 to induce DNA fragmentati on downstream Of JNK activation. (C) 1999 Academic Press.