Cell spreading in Colo 201 by staurosporin is alpha 3 beta 1 integrin-mediated with tyrosine phosphorylation of Src and tensin.
Staurosporin, a broad-spectrum kinase inhibitor, induced cell spreading in a human colon cancer cell line, Colo 201. On collagen and laminin, cell spreading was induced in more than 90% of the cells and was dependent on very late activation antigen-3, as shown by an antibody inhibition assay. Cell spreading required divalent cations and showed the order of preference Mn2+ > Mg2+ > Ca2+. On fibronectin, only about 30% of the cells were observed to spread, and spreading occurred via a non-integrin, RGD-independent pathway. Staurosporin-induced spreading was inhibited by treatment with tyrosine kinase inhibitors herbimycin A and methyl 2,5-dihydroxycinnamate. Despite the presence of staurosporin, seven proteins (220, 175, 150, 98, 62, 58, and 45 kDa) showed increased levels of tyrosine phosphorylation in association with cell adhesion. Two of these (58 and 220 kDa) were identified by immunoprecipitation as Src product and tensin, respectively. Flow cytometric analysis showed that the Colo 201 cells expressed the alpha 2, alpha 3, alpha 6, and beta 1 chains of integrin, but expression of these chains was not influenced by staurosporin. Immunofluorescence microscopy revealed that the alpha 3 chain, diffusely expressed on the cell surface in the absence of staurosporin, was concentrated at focal adhesion plaques after staurosporin treatment. Neither alpha 2 nor alpha 6 was focalized by the treatment.
Yoshimura
M
M
Department of Biochemistry, Osaka University Medical School, Japan.
Nishikawa
A
A
Nishiura
T
T
Ihara
Y
Y
Kanayama
Y
Y
Matsuzawa
Y
Y
Taniguchi
N
N
D000470:Alkaloids / Q000494:pharmacology*; D000595:Amino Acid Sequence; D002413:Cations, Divalent; D002448:Cell Adhesion / Q000187:drug effects*; D005109:Extracellular Matrix / Q000378:metabolism; D066298:In Vitro Techniques; D016023:Integrins / Q000502:physiology*; D008840:Microfilament Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D009842:Oligopeptides; D019000:Phosphotyrosine; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism*; D016029:Receptors, Very Late Antigen / Q000502:physiology*; D019311:Staurosporine; D000070876:Tensins; D014407:Tumor Cells, Cultured; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000378:metabolism
D000470:Alkaloids; D002413:Cations, Divalent; D016023:Integrins; D008840:Microfilament Proteins; D009842:Oligopeptides; D016029:Receptors, Very Late Antigen; D000070876:Tensins; D019000:Phosphotyrosine; D014443:Tyrosine; C047981:arginyl-glycyl-aspartic acid; D011505:Protein-Tyrosine Kinases; D016392:Proto-Oncogene Proteins pp60(c-src); D019311:Staurosporine