Murine colonic mucosa hyperproliferation. II. PKC-β activation and cPKC-mediated cellular CFTR overexpression

Abstract

In the companion article (Umar S, Scott J, Sellin JH, Dubinsky WP, and Morris AP, Am J Physiol Gastrointest Liver Physiol 278: 753–764, 2000), we have shown that transmissible murine colonic hyperplasia (TMCH) increased cellular cystic fibrosis transmembrane conductance regulator (CFTR) mRNA and protein expression, relocalized CFTR within colonocytes, and enhanced mucosal cAMP-dependent Cl−secretion. We show here that these changes were dependent on elevated cellular levels of membrane-bound Ca2+- and diacylglycerol-sensitive protein kinase C (PKC) activity (12-fold), induced by selective (3- to 4-fold) rises in conventional PKC (cPKC) isoform expression and membrane translocation. Three cPKC isoforms were detected in isolated crypts: α, β1, and β2. cPKC-β1 rises preceded and those of cPKC-α and cPKC-β2 paralleled cellular hyperproliferation and its effects on CFTR expression and cAMP-dependent Cl−current secretion. Only cPKC-β1 and cPKC-β2 were membrane translocated during TMCH. Furthermore, only cPKC-β1 trafficked to the nucleus, whereas cPKC-β2 remained partitioned among cytosolic, membrane, and cytoskeletal subcellular fractions. Modest increases in novel PKC-ε (nPKC-ε) expression and subcellular membrane partitioning were recorded during TMCH, but no changes were seen for PKC-δ or -η. No nPKC isoform nuclear partitioning was detected. The orally bioactive cPKC inhibitor Ro-32–0432 reversed both TMCH and elevated cellular CFTR mRNA levels, whereas a pharmacologically inert analog (Ro-31–6045) failed to inhibit either response. On the basis of these facts, we present a new hypothesis whereby PKC-dependent cellular proliferation promotes endogenous cellular CFTR levels. PKC-β1 was identified as a candidate regulatory PKC isoform.