Tissue and cellular distribution of the extended family of protein kinase C isoenzymes

Abstract

Polyclonal isoenzyme-specific antisera were developed against four calcium-independent protein kinase C (PKC) isoenzymes (δ, ∈, ∈′, and ζ) as well as the calcium-dependent isoforms (α, βI,βII, and γ). These antisera showed high specificities, high titers, and high binding affinities (3-370 nM) for the peptide antigens to which they were raised. Each antiserum detected a species of the predicted molecular weight by Western blot that could be blocked with the immunizing peptide. PKC was sequentially purified from rat brain, and the calcium-dependent forms were finally resolved by hydroxyapatite chromatography. Peak I reacted exclusively with antisera to PKCγ, peak II with PKCβI and -βII, and peak III with PKCα. These same fractions, however, were devoid of immunoreactivity for the calcium-independent isoenzymes. The PKC isoenzymes demonstrated a distinclive tissue distribution when evaluated by Western blot and immunocytochemistry. PCKδ was present in brain, heart, spleen, lung, liver, ovary, pancreas, and adrenal tissues. PKC∈ was present in brain, kidney, and pancreas, whereas PKC∈′ was present predominantly in brain. PKCζ was present in most tissues, particularly the lung, brain, and liver. Both PKCδ and PKCf showed some heterogeneity of size among the different tissues. PKCα was present in all organs and tissues examined. PKCβI and -βII were present in greatest amount in brain and spleen. Although the brain contained the most PKCγ immunoreactivity, some immunostaining was also seen in adrenal tissue. These studies provide the first evidence of selective organ and tissue distributions of the calciumindependent PKC isoenzymes.