Murine colonic mucosa hyperproliferation. II. PKC-beta activation and cPKC-mediated cellular CFTR overexpression.
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 Ca(2+)- 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: alpha, beta1, and beta2. cPKC-beta1 rises preceded and those of cPKC-alpha and cPKC-beta2 paralleled cellular hyperproliferation and its effects on CFTR expression and cAMP-dependent Cl(-) current secretion. Only cPKC-beta1 and cPKC-beta2 were membrane translocated during TMCH. Furthermore, only cPKC-beta1 trafficked to the nucleus, whereas cPKC-beta2 remained partitioned among cytosolic, membrane, and cytoskeletal subcellular fractions. Modest increases in novel PKC-epsilon (nPKC-epsilon) expression and subcellular membrane partitioning were recorded during TMCH, but no changes were seen for PKC-delta or -eta. 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-beta1 was identified as a candidate regulatory PKC isoform.
Umar
S
S
Department of Integrative Biology, Pharmacology, and Physiology, Division of Gastroenterology, Hepatology, and Nutrition, The University of Texas Health Science Center at Houston, Medical School, Houston 77030, USA.
Sellin
J H
JH
Morris
A P
AP
D000818:Animals; D002455:Cell Division / Q000187:drug effects; D002462:Cell Membrane / Q000201:enzymology; D003106:Colon / Q000473:pathology / Q000503:physiopathology*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics*; D003599:Cytoskeleton / Q000201:enzymology; D003600:Cytosol / Q000201:enzymology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D006965:Hyperplasia; D007211:Indoles / Q000494:pharmacology; D007413:Intestinal Mucosa / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology*; D007700:Kinetics; D008297:Male; D051379:Mice; D011493:Protein Kinase C / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D011758:Pyrroles / Q000494:pharmacology
D004791:Enzyme Inhibitors; D007211:Indoles; D011518:Proto-Oncogene Proteins; D011758:Pyrroles; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; C085989:Ro 32-0432; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D011493:Protein Kinase C