SYNOPSIS Protein kinase C (PKC) has been in the limelight since the discovery three decades ago that it acts as a major receptor for the tumor-promoting phorbol esters. Phorbol esters, with their potent ability to activate two of the three classes of PKC isozymes, have remained the best pharmacological tool for directly modulating PKC activity. However, with the discovery of other phorbol ester-responsive proteins, the advent of various small-molecule and peptide modulators, and the need to distinguish isozyme-specific activity, the pharmacology of PKC has become increasingly complex. Not surprisingly, many of the compounds originally touted as direct modulators of PKC have subsequently been shown to hit many other cellular targets and, in some cases, not even directly modulate PKC. The complexities and reversals in PKC pharmacology have led to widespread confusion about the current status of the pharmacological tools available to control PKC activity. Here, we aim to clarify the cacophony in the literature regarding the current state of bona fide and discredited cellular PKC modulators, including activators, small-molecule inhibitors, and peptides, and also address the use of genetically-encoded reporters and of PKC mutants to measure the effects of these drugs on the spatiotemporal dynamics of signaling by specific isozymes.

Background: Based on their in vitro effects, ZIP and chelerythrine have been used as PKM inhibitors and staurosporine as a negative control to implicate PKM in memory. Results: ZIP and chelerythrine do not and staurosporine does inhibit PKM in cells and brain slices. Conclusion: Cellular pharmacology of PKM contrasts with its in vitro profile. Significance: Contrary to current dogma, PKM may not mediate memory.

Background: Connexin43, a ubiquitous gap junction protein, is phosphorylated by protein kinase C on serine 368. Results: After PKC␦ activation, phospho-Ser-368 Connexin43 channels segregated into the gap junction center and were subsequently internalized and degraded. Conclusion: PKC␦ phosphorylation triggered internalization and degradation of Connexin43 channels without dephosphorylation. Significance: Differential phosphorylation events are used to sort and traffic Connexin43 channels within gap junctions and into the cytoplasm.

Background: PKC␦ signaling to mitochondria affects cellular apoptosis and metabolism. Results: A structure-function study using FRET-based imaging reveals that PKC␦ binds to and is active at mitochondria via multiple isozyme-specific determinants. Conclusion: Determinants unique to PKC␦ drive an interaction with mitochondria distinct from canonical PKC translocation to membranes. Significance: Isozyme-specific interaction of PKC␦ with mitochondria constitutes a novel recruitment mechanism and increases mitochondrial respiration.

Extended abstract of a paper presented at Microscopy and Microanalysis 2012 in Phoenix, Arizona, USA, July 29 – August 2, 2012.