title: Binding of the renal epithelial cell line LLC-PK1 to laminin is regulated by protein kinase C.
abstract: The alpha6beta1 integrin heterodimer has been implicated in the mediation of renal epithelial cell binding to laminin, and it has been suggested that this binding is important for renal morphogenesis and development. Studies of nonrenal cells have suggested that the functional activity of alpha6beta1 integrin is regulated by protein kinase C (PKC) activity. In this study, the binding of a renal epithelial cell line, LLC-PK1, to laminin was characterized and the role of PKC activity in the modulation of binding was investigated. LLC-PK1 cells bound to laminin-coated surfaces in a time- and laminin concentration-dependent manner. Binding was strongly inhibited by anti-beta1 integrin antibodies and by anti-alpha6 integrin antibodies. Antibodies against alpha2 integrin and a3 integrin had little inhibitory effect. Cells bound to both whole laminin and laminin fragment E8, i.e., the fragment to which the alpha6beta1 integrin heterodimer binds. Exposure of cells to PKC activators for as little as 2 h enhanced cell binding to laminin approximately twofold, in a protein synthesis-dependent manner. PKC inhibitors antagonized this effect. PKC-stimulated binding was also inhibited by anti-beta1 integrin and anti-alpha6 integrin antibodies. PKC activation did not alter expression of beta1 integrin subunits at the cell surface after short time periods (2 to 4 h), but expression was increased after longer time periods (24 h). These results indicate that the renal epithelial cell line LLC-PK1 binds to laminin via the alpha6betal integrin heterodimer and binding is enhanced by PKC activation. The PKC-mediated enhancement of binding requires protein synthesis and is mediated in part by activation of surface alpha6beta1 integrin.
authors.affiliation: Department of Physiology and Biophysics, University of Medicine and Dentistry of New Jersey, Robert Wood Johnson Medical School, Piscataway 08854-5635, USA.
authors.forename: R
authors.initials: R
authors.lastname: Kapadia
authors.forename: P D
authors.initials: PD
authors.lastname: Yurchenco
authors.forename: K
authors.initials: K
authors.lastname: Amsler
mesh_terms: D000818:Animals; D001665:Binding Sites; D004305:Dose-Response Relationship, Drug; D016023:Integrins / Q000187:drug effects / Q000378:metabolism; D007668:Kidney / Q000166:cytology; D018374:LLC-PK1 Cells / Q000187:drug effects / Q000378:metabolism*; D007797:Laminin / Q000378:metabolism* / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects* / Q000502:physiology; D011493:Protein Kinase C / Q000378:metabolism*; D012016:Reference Values; D013552:Swine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
chemical_list: D016023:Integrins; D007797:Laminin; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate