Inhibitory effect of regucalcin on Ca2+/calmodulin-dependent cyclic AMP phosphodiesterase activity in rat kidney cytosol

Abstract

The effect of regucalcin, a novel Ca2+-binding protein, on Ca2+/calmodulin-dependent cyclic adenosine monophosphate (AMP) phosphodiesterase activity in the cytosol of rat renal cortex was investigated. Regucalcin with physiologic concentration (10[-7] M) in rat kidney had no effect on cyclic AMP phosphodiesterase activity in the absence of CaCl2 and calmodulin. However, the activatory effect of both CaCl2 (10 microM) and calmodulin (20 U/ml) on cyclic AMP phosphodiesterase was markedly inhibited by the addition of regucalcin (10[-8] to 10[-6] M) in the enzyme reaction mixture. The inhibitory effect of regucalcin on the enzyme activity was also seen in the presence of CaCl2 (5-50 microM) or calmodulin (5-50 U/ml) with increasing concentrations. The presence of trifluoperazine (10 microM), an antagonist of calmodulin, caused a partial inhibition of Ca2+/calmodulin-dependent cyclic AMP phosphodiesterase activity. This inhibition was further enhanced by the addition of regucalcin (10[-7] M). The inhibitory effect of regucalcin (10[-7] M) was not seen in the presence of 20 microM trifluoperazine. Moreover, the activatory effect of calmodulin (20 U/ml) on cyclic AMP phosphodiesterase was not entirely seen, when calmodulin was added 10 min after incubation in the presence of CaCl2 (10 microM) and regucalcin (10[-7] M). The present results demonstrates that regucalcin has an inhibitory effect on Ca2+/calmodulin-dependent cyclic AMP phosphodiesterase activation in the cytosol of rat renal cortex.