Pig skin epidermal calmodulin: effect on calmodulin deficient phosphodiesterase

Abstract

Calmodulin, a calcium-dependent modulator protein, is known to mediate a great number of Ca++-dependent processes in various tissues. Although it was originally described as a protein activator of cyclic nucleotide phosphodiesterase, the sensitivity of phosphodiesterases to this compound are suggested to be variable from tissue to tissue. In order to determine whether there was calmodulin-like activity in pig skin epidermis and to see its relationship to epidermal phosphodiesterase, we used an established calmodulin deficient phosphodiesterase system prepared from bovine heart. Calmodulin deficient phosphodiesterase prepared from bovine heart was markedly stimulated by the addition of pig skin (epidermal) boiled extract in the presence of calcium. Boiled skin extract alone had only little phosphodiesterase activity by itself. This effect of boiled skin extract on bovine heart phosphodiesterase was inhibited by the addition of EGTA, a divalent metal ion chelator of relative Ca++ specificity. At a fixed concentration of EGTA, increasing the Ca++ concentration counteracted the effect of EGTA. Pure pig skin epidermis (separated by trypsinization, NaBr, CaCl2-sucrose or NH4Cl treatment) was also shown to have heat-stable calmodulin activity. In contrast to the bovine heart phosphodiesterase, epidermal phosphodiesterase was only partially inhibited when Ca++ was removed by EGTA. The addition of boiled skin extract on the crude extract of epidermal phosphodiesterase had minimal effect on the enzyme activity. Overall results indicate that although pig skin epidermis contains significant amount of calmodulin, the regulation of phosphodiesterase may not be the main biological activity of epidermal calmodulin.