title: Regulation of SPIN90 phosphorylation and interaction with Nck by ERK and cell adhesion.
abstract: SPIN90 is a widely expressed Nck-binding protein that contains one Src homology 3 (SH3) domain, three Pro-rich motifs, and a serine/threonine-rich region, and is known to participate in sarcomere assembly during cardiac myocyte differentiation. We used in vitro binding assays and yeast two-hybrid screening analysis to identify Nck, betaPIX, Wiscott-Aldrich syndrome protein (WASP), and ERK1 as SPIN90-binding proteins. It appears that betaPIX, WASP, and SPIN90 form a complex that interacts with Nck in a manner dependent upon cell adhesion to extracellular matrix. The betaPIX.WASP.SPIN90.Nck interaction was abolished in suspended and cytochalasin D-treated cells, but was recovered when cells were replated on fibronectin-coated dishes. The SPIN90.betaPIX.WASP complex was stable, even in suspended cells, suggesting SPIN90 serves as an adaptor molecule to recruit other proteins to Nck at focal adhesions. In addition, we found that overexpression of the SPIN90 SH3 domain or Pro-rich region, respectively, abolished SPIN90.Nck and SPIN90.betaPIX interactions, resulting in detachment of cells from extracellular matrix. SPIN90 was phosphorylated by ERK1, which was, itself, activated by cell adhesion and platelet-derived growth factor. Such phosphorylation of SPIN90 likely promotes the interaction of the SPIN90.betaPIX.WASP complex and Nck. It thus appears that the interaction of the betaPIX.WASP.SPIN90 complex with Nck is crucial for stable cell adhesion and can be dynamically modulated by SPIN90 phosphorylation that is dependent on cell adhesion and ERK activation.
authors.affiliation: Department of Life Science, Kwangju Institute of Science and Technology, 1 Oryong-dong, Buk-gu, Gwangju 500-712, Korea.
authors.forename: Chol Seung
authors.initials: CS
authors.lastname: Lim
authors.forename: Sung Hyun
authors.initials: SH
authors.lastname: Kim
authors.forename: Jin Gyoung
authors.initials: JG
authors.lastname: Jung
authors.forename: Jin-Kyu
authors.initials: JK
authors.lastname: Kim
authors.forename: Woo Keun
authors.initials: WK
authors.lastname: Song
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D020816:Amino Acid Motifs; D002448:Cell Adhesion; D018797:Cell Cycle Proteins / Q000378:metabolism; D002454:Cell Differentiation; D015638:Cytochalasin D / Q000494:pharmacology; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005109:Extracellular Matrix / Q000378:metabolism; D005353:Fibronectins / Q000378:metabolism; D022001:Focal Adhesions; D005982:Glutathione Transferase / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D008957:Models, Genetic; D032342:Muscle Cells / Q000378:metabolism; D009124:Muscle Proteins / Q000378:metabolism*; D015513:Oncogene Proteins / Q000378:metabolism*; D010766:Phosphorylation; D011233:Precipitin Tests; D011392:Proline / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000378:metabolism; D064067:Rho Guanine Nucleotide Exchange Factors; D015398:Signal Transduction; D014162:Transfection; D020798:Two-Hybrid System Techniques; D051316:Wiskott-Aldrich Syndrome Protein; D018909:src Homology Domains
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins; D018076:DNA, Complementary; D004791:Enzyme Inhibitors; D005353:Fibronectins; D020662:Guanine Nucleotide Exchange Factors; D009124:Muscle Proteins; C486062:NCKIPSD protein, human; C078332:Nck protein; D015513:Oncogene Proteins; D011506:Proteins; D064067:Rho Guanine Nucleotide Exchange Factors; C495767:WAS protein, human; D051316:Wiskott-Aldrich Syndrome Protein; D015638:Cytochalasin D; D011392:Proline; D005982:Glutathione Transferase; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases