pp125FAK in human melanocytes and melanoma: expression and phosphorylation.
Focal adhesion kinase (pp125FAK) is a nonreceptor tyrosine kinase which colocalizes with integrins to focal contacts, sites where multiple proteins interact to regulate the assembly of the actin cytoskeleton. Autophosphorylation and activation of pp125FAK occur after integrin clustering or cell adhesion to ligands through cognate integrin receptors and are postulated to mediate integrin signaling events. In this report we examined pp125FAK expression and phosphorylation in normal human melanocytes, an adherent human metastatic melanoma cell line (SKMEL28), and a nonadherent human metastatic melanoma cell line (SKMEL1). We show that SKMEL28 cells express constitutively phosphorylated pp125FAK and that pp125FAK phosphorylation in melanocytes is induced by phorbol esters and growth factors present in melanocyte growth medium. Focal adhesion kinase phosphorylation could be enhanced by b1 integrin-activating antibodies in human melanocytes, but not in SKMEL28 cells. In contrast with SKMEL28 cells, constitutive phosphorylation of pp125FAK was not observed in SKMEL1 cells, and incubation with activating b1 integrin antibodies had no effect on pp125FAK phosphorylation. Absence of pp125FAK phosphorylation in SKMEL1 cells was not due to lack of expression of pp125FAK, as shown by immunoprecipitation of the pp125FAK protein from cell lysates. However, b1 integrin expression was significantly less in SKMEL1 cells than in human melanocytes and SKMEL28 cells. This study further supports the importance of integrins in pp125FAK-mediated signaling and indicates that transformation-related changes in pp125FAK phosphorylation exist in human melanocytes and melanoma cells.
Scott
G
G
Department of Dermatology, University of Rochester School of Medicine and Dentistry, New York 14642, USA.
Liang
H
H
D000906:Antibodies / Q000494:pharmacology; D015153:Blotting, Western; D015815:Cell Adhesion Molecules / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D015870:Gene Expression; D006801:Humans; D019012:Integrin beta1; D016023:Integrins / Q000276:immunology / Q000502:physiology; D046911:Macromolecular Substances; D008544:Melanocytes / Q000378:metabolism*; D008545:Melanoma / Q000378:metabolism*; D010766:Phosphorylation; D019000:Phosphotyrosine; D011505:Protein-Tyrosine Kinases / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D011972:Receptor, Insulin / Q000378:metabolism; D014407:Tumor Cells, Cultured; D014443:Tyrosine / Q000031:analogs & derivatives / Q000032:analysis
D000906:Antibodies; D015815:Cell Adhesion Molecules; D019012:Integrin beta1; D016023:Integrins; D046911:Macromolecular Substances; D019000:Phosphotyrosine; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human