Direct Activation of Protein Kinase C by 1α,25-Dihydroxyvitamin D3

Abstract

The key metabolite of vitamin D3, 1 alpha,25-dihydroxyvitamin D3 (1,25-D3), induces rapid cellular responses that constitute a so-called "non-genomic" response. This effect is distinguished from its "classic" genomic role in calcium homeostasis involving the nuclear 1,25-D3 receptor. Evidence is presented that protein kinase C (PKC) is directly activated by 1,25-D3 at physiological concentrations (EC50 = 16 +/- 1 nM). The effect was demonstrable with single PKC-alpha, -gamma, and -epsilon isoform preparations, assayed in a system containing only purified enzyme, substrate, co-factors, and lipid vesicles, from which it is inferred that a direct interaction with the enzyme is involved. The finding that calcium-independent isoform PKC-epsilon was also activated by 1,25-D3 shows that the calcium binding C2 domain is not required. The level of 1,25-D3-induced activation, paired with either diacylglycerol or 4 beta-12-O-tetradecanoylphorbol-13-acetate, was greater than that achievable by any individual activator alone, each at a saturating concentration, a result that implies two distinct activator sites on the PKC molecule. Phosphatidylethanolamine present in the lipid vesicles potentiated 4 beta-12-O-tetradecanoylphorbol-13-acetate- and diacylglycerol-induced PKC activities, whereas 1,25-D3-induced activity decreased, consistent with 1,25-D3-activated PKC possessing a distinct conformation. The results suggest that PKC is a "membrane-bound receptor" for 1,25-D3 and that it could be important in the control of non-genomic cellular responses to the hormone.

Bryostatin-1 and 1α,25-dihydroxyvitamin D3 synergistically stimulate the differentiation of NB4 acute promyelocytic leukemia cells

“…It has been demonstrated that 1␣,25(OH) 2 D 3 alone can directly activate PKC and increase VDR phosphorylation. 35,36 PMA was reported to up-regulate VDR mRNA and the VDR protein levels in myeloid cells or non-myeloid cells by activation of PKC. 37,38 Furthermore, it has been suggested that PKC down-regulation rather than activation by bryostatin or PMA plays a role in the differentiation of several cell lines.…”

Mechanism of Diacylglycerol-induced Membrane Targeting and Activation of Protein Kinase Cδ

“…As shown in Table III, the C1A domain bound DiC 8 with a K d of 85 nM, whereas the C1B domain showed no detectable binding. Thus, as was the case with PKC␣ (38,74), the C1A and C1B domains of PKC␦ have opposite affinities for DAG and phorbol ester, i.e. the C1A domain with high affinity for DAG and the C1B domain with high affinity for phorbol ester.…”

Inhibition of Membrane Lipid-independent Protein Kinase Cα Activity by Phorbol Esters, Diacylglycerols, and Bryostatin-1

“…However, in recent studies of activator binding to PKC␣, using the fluorescent phorbol ester sapintoxin D (SAPD), we showed that diacylglycerols and phorbol esters interact with differing affinities with two activator binding sites on the enzyme molecule (34 -36). Furthermore, it was shown that the site with low affinity for phorbol esters binds diacylglycerol with a relatively higher affinity, suggesting that the specificity of this site may differ from the high affinity phorbol ester-binding site (35). Interaction of diacylglycerol with this low affinity phorbol ester-binding site was found to lead to an enhancement in the level of high affinity phorbol ester binding and consequently to a potentiated level of PKC activity.…”