Evidence for a role of conventional protein kinase-C alpha in the control of homotypic contacts and cell scattering of HT-29 human intestinal cells.
Incubation of HT-29 M6 cells with the phorbol ester phorbol 12-myristate 13-acetate (PMA) induces cell scattering, loss of cellular contacts and inactivation of E-cadherin. We have investigated the involvement of different protein kinase C (PK-C) isoforms in these processes using specific activators. Thymeleatoxin, a derivative of mezerein that activates conventional PK-Cs (cPK-Cs) but not novel PK-Cs (nPK-Cs), promoted effects that were similar to those of PMA, i.e. at concentrations of 200 nM it induced scattering of HT-29 M6 colonies, loss of homotypic contacts and dissociation of E-cadherin from the cytoskeleton. Among the isoforms activated by this compound, only cPK-C alpha was detected in HT-29 M6 cells by Western blot. The specificity of this compound with respect to the rest of the PK-C isoforms present in these cells was determined; thymeleatoxin induced, as did PMA, the translocation of cPK-C alpha from the cytosol to the membrane and the cytoskeleton, and its partial down-regulation. On the other hand, thymeleatoxin did not modify the cellular levels or localization of nPK-C epsilon or atypical PK-C zeta. "In vitro' assays also showed that thymeleatoxin did not activate nPK-C epsilon at the concentrations added to the cell cultures. These results indicate that thymeleatoxin is selective for cPK-C alpha over nPK-C epsilon and show a role for the former enzyme in the regulation of cell-cell contacts and the inactivation of E-cadherin in HT-29 M6 cells.
Llosas
M D
MD
Department d'Immunologia, Institut Municipal d'Investigació Mèdica, Universitat Autònoma de Barcelona, Spain.
Batlle
E
E
Coll
O
O
Skoudy
A
A
Fabre
M
M
García de Herreros
A
A
D000595:Amino Acid Sequence; D000906:Antibodies / Q000494:pharmacology; D015820:Cadherins / Q000378:metabolism; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology*; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology*; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000037:antagonists & inhibitors / Q000235:genetics / Q000276:immunology; D010703:Phorbol Esters / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D051571:Protein Kinase C-alpha; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D000906:Antibodies; D015820:Cadherins; D007527:Isoenzymes; D010446:Peptide Fragments; D010703:Phorbol Esters; C080287:thymeleatoxin; C496628:PRKCA protein, human; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D013755:Tetradecanoylphorbol Acetate