Ca /calmodulin-dependent protein kinase II inhibition by heparin in mesangial cells

Xiao, Weiqun, Ying Liu, and Douglas M. Templeton. Ca / calmodulin-dependent protein kinase II inhibition by heparin in mesangial cells. Am J Physiol Renal Physiol 288: F142–F149, 2005. First published September 21, 2004; doi:10.1152/ajprenal.00145.2004.— Heparin exerts an antiproliferative effect in smooth muscle cells, and the Ca /calmodulin-dependent protein kinase (CaMK) signaling pathway is heparin sensitive. Here, we report that transfection with a truncated 326-amino acid fragment of CaMK-II increases basal activity of CaMK-II in mesangial cells. Ionomycin increased CaMK-II activity in both transfected and untransfected cells, with a concomitant increase in activated Ca /calmodulin. Heparin (1 g/ ml), but not chondroitin or dermatan sulfate, significantly attenuated both serumor ionomycin-induced CaMK-II activity, and attendant c-fos mRNA expression, but did not affect upstream Ca /calmodulin. Autophosphorylation of Thr286 generates an autonomously active CaMK-II. Both serum and ionomycin increased phosphorylation at this site and increased CaMK-II activity in antiphosphothreonine immunoprecipitates. Heparin (1 g/ml) did not inhibit phosphorylation of Thr286 (although much higher concentrations did). Replacement of Thr286 with Asp produces a constitutively active mutant that was insensitive to ionomycin but was inhibited by heparin maximally at 1 g/ml. These results suggest that heparin at physiological concentrations acts at or downstream of CaMK-II to suppress its activity independent of an effect on autophosphorylation.