Protein kinase C isoforms in pituitary cells displaying differential sensitivity to phorbol ester

Investigations with protein kinase C (PKC) isoform-specific antisera, revealed distinct profiles of PKC isoform content amongst pituitary tissues. Western analysis revealed the alpha, beta, delta, epsilon, zeta and theta isoforms of PKC are present in rat anterior and posterior pituitary tissue as well as in the GH(3) somatomammotrophic cell line. AtT-20/D16-V corticotrophic and alpha T3-1 gonadotrophic murine cell lines contained no PKC-delta. The gamma or eta isoforms were undetected in any pituitary tissue. PKC activity measurements revealed Ca2+-independent PKCs in alpha T3-1 and GH(3) cells which were more sensitive to activation by phorbol-dibutyrate (PDBu) than the corresponding PKC activity found in COS cells. However, Ca2+-dependent PKC activities were of similar sensitivity to PDBu in GH(3), alpha T3-1 and COS cells, indicating that functional differences observed in PDBu-sensitivity in these cells may be due to differential activation of Ca2+-independent PKC isoforms. Moreover, substrate-specificity of these PKCs were also compared indicating that the amount of Ca2+-dependency of the observed PKC activity from the same pituitary tissue is dependent upon the substrate utilized by the PKC isotypes present. These findings explain differential sensitivities of PKC-mediated actions that have previously been observed in a range of pituitary cells.