title: Involvement of protein kinase C in translocation of desmoplakins from cytosol to plasma membrane during desmosome formation in human squamous cell carcinoma cells grown in low to normal calcium concentration.
abstract: The intracellular signal transduction mechanism leading to desmosome formation in low-calcium-grown keratinocytes after addition of calcium to the medium was studied by immunofluorescence using antibodies to desmoplakins I and II (cytoplasmic desmosomal proteins) and by electron microscopy before and after addition of calcium; protein kinase C (PKC) activators 12-O-tetradecanoylphorbol-13-acetate (TPA), phorbol-12,13-dibutyrate (PDBu), and 1,2-dioctanoylglycerol (DOG); calcium ionophore A23187; selective PKC inhibitors 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H-7) and staurosporine; and a Ca2+/calmodulin-dependent kinase inhibitor, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7). In previous studies using a low-calcium-grown human epidermal squamous cell carcinoma, we have shown that an increase in extracellular Ca2+ caused a four-fold increase in PKC activity and addition of TPA (10 ng/ml) induced a transient increase in membrane-bound PKC activity in association with cell-cell contact formation. The present study showed that TPA (10 ng/ml). PDBu (10 ng/ml), and DOG (1 mg/ml) induced a rapid cell-cell contact and redistribution of desmoplakins from cytoplasm to the plasma membrane with desmosome formation within 60-120 min, which was similar, although less marked, to the effect of increased Ca2+. The TPA-induced desmosome formation was inhibited by selective PKC inhibitors, H-7 (20 microM) or staurosporine (100 nM). On the other hand, calcium ionophore A23187 induced only a temporary increase in the number of desmoplakin-containing fluorescent spots in the cytoplasm and a temporary cell-cell attachment without desmosome formation. The calcium-induced desmosome formation was partially inhibited by 20-100 microM H-7 or 100 nM staurosporine; however, it was not inhibited by W-7 at a concentration of 25 microM, at which this agent selectively inhibits calmodulin-dependent protein kinase. These results suggest that PKC activation plays an important role in desmoplakin translocation from the cytoplasm to the plasma membrane as one of the processes of calcium-induced desmosome formation.
authors.affiliation: Department of Dermatology, Jichi Medical School, Tochigi, Japan.
authors.forename: H M
authors.initials: HM
authors.lastname: Sheu
authors.forename: Y
authors.initials: Y
authors.lastname: Kitajima
authors.forename: H
authors.initials: H
authors.lastname: Yaoita
mesh_terms: D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology*; D002294:Carcinoma, Squamous Cell / Q000473:pathology* / Q000503:physiopathology / Q000648:ultrastructure; D002462:Cell Membrane / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000276:immunology / Q000378:metabolism / Q000493:pharmacokinetics*; D003600:Cytosol / Q000378:metabolism*; D051180:Desmoplakins; D003896:Desmosomes / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D004075:Diglycerides / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D006801:Humans; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism / Q000502:physiology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured / Q000473:pathology / Q000648:ultrastructure
chemical_list: D003598:Cytoskeletal Proteins; C495294:DSP protein, human; D051180:Desmoplakins; D004075:Diglycerides; C044295:1,2-dioctanoylglycerol; D015240:Phorbol 12,13-Dibutyrate; D000001:Calcimycin; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium