Differential localization of protein kinase C delta by phorbol esters and related compounds using a fusion protein with green fluorescent protein

Enzyme localization often plays a controlling role in determining its activ ity and specificity. Protein kinase C (PKC) has long been known to transloc ate in response to physiological stimuli as well as to exogenous ligands su ch as the phorbol esters, We report here that different phorbol derivatives and related ligands, selected for differences in chemical structure and pr ofile of biological activity, induce distinct patterns of redistribution of PKC delta, Localization of a PKC delta-green fluorescent protein (GFP) fus ion construct was monitored in living Chinese hamster ovary cells as a func tion of ligand, concentration, and time using confocal laser scanning micro scopy, delta-PKC-GFP was expressed predominantly in the cytoplasm, with som e in the nucleus and perinuclear region. Phorbol 12-myristate 13-acetate (P MA) induced plasma membrane translocation followed by slower nuclear membra ne translocation. As the concentration of PMA increased, the proportion of nuclear to plasma membrane localization increased markedly. In contrast to PMA, bryostatin 1, a unique activator of PKC that induces a subset of PIMA mediated responses while antagonizing others, at all doses induced almost e xclusively nuclear membrane translocation, Like PMA, the complete tumor pro moter 12-deoxyphorbol 13-tetradecanoate induced plasma membrane and slower nuclear membrane translocation, whereas the inhibitor of tumor promotion 12 -deoxyphorbol 13-phenylacetate, which differs only in its side chain, induc ed a distinctive distribution of PKC delta-GFP, Finally, the novel constrai ned diacylglycerol derivative B8-DL-B8 induced a slow Golgi localization. W e speculate that differential control of PRC delta localization may provide an interesting strategy for producing ligands with differential biological consequences.