Role and Regulation of Protein Kinase C Isoenzymes

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DESCRIPTION (provided by applicant): Conceptually, Protein kinase C (PKC) has served as the prototype of lipid-regulated proteins, best illustrated in the case of the phosphatidylinositol (PI) cycle whereby PI-derived diacylglycerol (DAG) serves as the direct activator of PKC. Given the rapid turnover of the PI cycle (within 90 seconds of receptor engagement), this paradigm of PKC activation has by necessity focused on the very acute phase of PKC regulation. Importantly, this paradigm does not address other mechanisms of DAG generation, most notably through phospholipase D (PLD), and it does not distinguish mechanisms for regulating activation of specific isoenzymes. Extensive preliminary results have generated exciting information on novel mechanisms and functions of PKC. Briefly, we find that i) sustained activation of PKC (30-60 min) induces the translocation of PKC alpha and betaII (but not the closely related PKC betaI) to a novel juxtanuclear/pericentriolar compartment; ii) this compartment has features of a previously-identified (but not well-appreciated) component of recycling endosomes; iii) PLD is required for this translocation; and iv) translocation of PKC to this juxtanuclear compartment is accompanied by re-localization/sequestration of some plasma membrane proteins (e.g. transferrin receptor and glut4) and lipids (e.g. GM1) to this compartment. Based on these observations, we hypothesize that stimulation of PLD results in sustained activation of PKC alpha and betaII and their translocation to a pericentriolar compartment. This mechanism may play a key role in the sequestration and availability of recycling components of the plasma membrane. This hypothesis will be investigated by pursuing the following specific aims 1) Define and establish a novel translocation of PKC; 2) Define mechanisms of regulation of juxtanuclear PKC and role of PLD; 3) Define role of PKC in sequestration of plasma membrane and/or recycling components. If correct, this novel translocation and function of PKC may be involved in multiple physiologic and pathophysiologic processes that depend on availability of key proteins and/or lipids at the PM, including diabetes, and cancer.