Reversing the Paradigm: Protein Kinase C as a Tumor Suppressor

Abstract

The discovery in the 1980s that protein kinase C (PKC) is a receptor for the tumor-promoting phorbol esters fueled the dogma that PKC is an oncoprotein. Yet 30+ years of clinical trials for cancer using PKC inhibitors not only failed, but in some instances worsened patient outcome. The recent analysis of cancer-associated mutations, from diverse cancers and throughout the PKC family, revealed that PKC isozymes are generally inactivated in cancer, supporting a tumor suppressive function. In keeping with a bona fide tumor suppressive role, germline causal loss-of-function (LOF) mutations in one isozyme have recently been identified in lymphoproliferative disorders. Thus, strategies in cancer treatment should focus on restoring rather than inhibiting PKC.

Keywords: LOF; PKC; diacylglycerol; phorbol esters; tumor suppressor.

Figure 1

Cartoon showing the multiple inputs regulating the function of a conventional PKC. Newly-synthesized PKC is processed by a series of ordered phosphorylations that depend on the binding of Hsp90 to a conserved PXXP motif in the kinase domain [74], the kinase complex mTORC2 [75], and PDK-1 (reviewed in [6]). Phosphorylation at three priming sites, the activation loop, the turn motif, and the hydrophobic motif, allow PKC to adopt an autonhibited conformation in which the Ca²⁺-sensing C2 domain (yellow) clamps the autoinhibitory pseudosubstrate segment (red) in the substrate-binding cavity of the kinase domain (cyan), and the diacyglycerol-sensing C1 domains (orange) become masked. Upon elevation of intracellular Ca²⁺, the C2 domain engages on the plasma membrane via Ca²⁺ coordination to anionic lipids and binding of plasma membrane-localized PIP₂ to a surface on the domain. This positions PKC to bind its membrane-embedded ligand, diacylglycerol (DG), an event that releases the pseudosubstrate for maximal activation of PKC. This active PKC phosphorylates downstream substrates, such as K-RAS, to suppress oncogenic signaling. The open conformation of PKC is sensitive to dephosphorylation, with the first event being dephosphorylation of the hydrophoboic motif catalyzed by PHLPP; subsequent dephosphorylation by PP2A produces a fully dephosphorylated PKC that is shunted for degradation by a proteosomal pathway. However, binding of HSP70 to the dephosphorylated turn motif allows PKC to become rephosphorylated to sustain the signaling lifetime of the enzyme. Phorbol esters (not shown) bind with two-orders of magnitude higher affinity than diacylglycerol (highlighted in yellow) to the C1B domain and are not readily metabolized, trapping PKC in the open, active conformation and resulting in chronic loss, or down-regulation, of PKC. Novel PKC isozymes are regulated by similar mechanisms except their C2 domain does not function as a Ca²⁺ sensor. Atypical PKC isozymes are activated upon binding to specific protein scaffolds that tether the pseudosubstrate out of the substrate-binding cavity. Proteins indicated in grey are key regulators of the steady-state levels of PKC: HSP70, HSP90, mTORC2, and PDK-1 function to increase the steady-state levels of PKC by permitting processing phosphorylations, and PHLPP and PP2A decrease the steady-state levels of PKC by catalyzing the dephosphorylation of PKC. Targeting any of these proteins in cancer treatments will disrupt the balance of PKC signaling.

Figure I