The stimulation of MAP kinase by 1,25(OH)2-vitamin D3 in skeletal muscle cells is mediated by protein kinase C and calcium

In previous work we have demostrated that the steroid hormone 1,25(OH)(2)-vitamin D-3 [1,25(OH)(2)D-3] stimulates in skeletal muscle cells the phosphorylation and activity of the extracellular signal-regulated mitogen-activated protein (MAP) kinase isoforms ERK1 and ERK2. In the present study we evaluated the involvement of Ca2+ and protein kinase C (PKC) on 1,25(OH)(2)D-3-induced activation of MAP kinase. The hormone response was found to depend on PKC stimulation since it was attenuated by the PKC inhibitors calphostin C (100 nM) and bisindolylmaleimide 1 (30 nM) and PKC downregulation by prolonged treatment with the phorbol ester TPA (1 muM). Removal of external Ca2+ chelation of intracellular Ca2+ with BAPTA (5 muM), inhibition of phosphoinositide phospholipase C (PLC) by neomycin. the calmodulin antagonist fluphenazine (50 muM) and the specific inhibitor of calmodulin kinase III KN-62 (10 muM). significantly decreased 1,25(OH)(2)D-3-activation of MAP kinase. In addition, the Ca2+-channel blocker verapamil (5 muM) suppressed hormone-induced MAP kinase activity in these cells. Furthermore, the Ca2+-mobilizing agent thapsipargin and the Ca2+-inophore A23187 paralleled the phosphorylation of MAP kinase observed with 1,25(OH)(2)D-3. Taken together. these results indicate that PKC and Ca2+ are two upstream activators mediating the effects of 1,25(OH)(2)D-3 on MAP kinase in skeletal muscle cells. (C) 2001 Elsevier Science Ireland Ltd. All rights reserved.