Protein kinase C as a possible receptor protein of tumor-promoting phorbol esters.

TLDR

A tumor‑promoting phorbol ester, [3H]phorbol‑12,13‑dibutyrate, binds to Ca²⁺‑activated, phospholipid‑dependent protein kinase C only when Ca²⁺ and phospholipid are present. Binding of the phorbol ester to protein kinase C is specifically blocked by diacylglycerol, an endogenous activator of the enzyme. The phorbol ester forms an active quaternary complex with Ca²⁺, phospholipid, and protein kinase C; phosphatidylserine is most effective, other tumor‑promoting phorbols compete, the dissociation constant is 8 nM—matching the activation constant—and Scatchard analysis indicates one promoter per kinase, supporting PKC as the phorbol‑ester receptor.

Abstract

A tumor-promoting phorbol ester, [3H]phorbol-12,13-dibutyrate, may bind to a homogeneous preparation of Ca2+-activated, phospholipid-dependent protein kinase (protein kinase C) in the simultaneous presence of Ca2+ and phospholipid. This tumor promoter does not bind simply to phospholipid nor to the enzyme per se irrespective of the presence and absence of Ca2+. All four components mentioned above appear to be bound together, and the quaternary complex thus produced is enzymatically fully active for protein phosphorylation. Phosphatidylserine is most effective. Various other phorbol derivatives which are active in tumor promotion compete with [3H]phorbol-12,13-dibutyrate for the binding, and an apparent dissociation binding constant of the tumor promoter is 8 nM. This value is identical with the activation constant for protein kinase C and remarkably similar to the dissociation binding constant that is described for intact cell surface receptors. The binding of the phorbol ester is prevented specifically by the addition of diacylglycerol, which serves as activator of protein kinase C under physiological conditions. Scatchard analysis suggests that one molecule of the tumor promoter may bind to every molecule of protein kinase C in the presence of Ca2+ and excess phospholipid. It is suggestive that protein kinase C is a phorbol ester-receptive protein, and the results presented seem to provide clues for clarifying the mechanism of tumor promotion.