The Role of Protein Kinase C in Gastrointestinal Function and Disease

Structure of PKC isoforms. The structure for classical (cPKC), novel (nPKC), and atypical (aPKC) isoforms is shown schematically. They are divided into their regulatory and catalytic domains as shown. The N-terminal pseudosubstrate (PSEUDO), the phophatidyl-L-serine (PS), and the ATP (C3) binding domains and the C4 catalytic domain are common to all 3 subfamilies.⁷⁰ The cPKC and the nPKCs both contain functional 1,2-sn-diacylglycerol (DAG) binding domains (C1) and the cPKC contains a Ca⁺²-binding domain that is nonfunctional and more amino terminal in the nPKCs and completely absent in the aPKCs.

Model of PKC activation. (1) Newly synthesized PKC (PKCβII depicted here) binds to the cytoskeleton in its “immature” form. (2 and 3) During maturation, PDK-1 phosphorylates at Thr500 of C1, which makes the activation loop accessible for binding of the pseudosubstrate to the catalytic domain. This permits autophosphorylation at Thr641 and Ser660. (3) Once the phosphorylation is complete, the “mature” inactive PKC can then bind to the plasma membrane (4) via weak interactions with negatively charged lipid head groups. This binding is transient, and the PKC fluctuates between membrane bound and a free cytosolic equilibrium. During agonist activation, phospholipase type C (PLC) hydrolyzes lipid, releasing Ca⁺² and membrane associated 1,2-sn-diacylglycerol (DAG). (5) Ca⁺² binding to the C2 domain tethers the PKC to the membrane, allowing it to scan for DAG. (6) Upon interaction with DAG, the C1 domain of PKC forms a strong bond with phosphatidyl-L-serine (PS) in the membrane, resulting in an active, open PKC enzyme that is able to phosphorylate its appropriate substrate.

Involvement of PKC and colorectal cancer. (A) β-Catenin regulation. β-Catenin is released into the cytosol where it is bound by adenomatous polyposis coli (APC) protein and phosphorylated by GSK-3β. These 2 events lead to the ubiquitination and degradation of β-catenin. Activation of cPKCβII results in phosphorylation of GSK-3β by PKCβII, and this inactivates GSK-3βm, resulting in cytosolic and eventually nuclear accumulation of β-catenin. Once in the nucleus, β-catenin acts as a transcriptional regulator leading to loss of cell regulation and eventually malignancy. (B) COX-2 induction in colonic epithelia. Over-expression of cPKCβII results in induction of COX-2 expression, which, in turn, down-regulates expression of the TGF-βRII. This results in hyperproliferation caused by a loss of TGF-β regulation, as well as increased proliferative signals mediated by COX-2 activity (ie, prostaglandin secretion). This activity of cPKCβII is down-regulated by nonfermentable brans, cellulose, and ω-3 polyunsaturated fatty acids (PUFA) and is augmented by azoxymethane (AOM). (C) COX-2 induction in colonic myofibroblasts. IL-1 binds to its receptor, activating aPKCζ and NF-κB. This activation of aPKCζ results in generation of reactive oxygen species (ROS) as well as translocation of aPKCζ into the nucleus. Through an unknown mechanism, aPKCζ and ROS potentiate COX-2 transcription.