CELL SPREADING IN COLO-201 BY STAUROSPORIN IS ALPHA-3-BETA-1-INTEGRINMEDIATED WITH TYROSINE PHOSPHORYLATION OF SRC AND TENSIN

Staurosporin, a broad-spectrum kinase inhibitor, induced cell spreadin g 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 dep endent on very late activation antigen-3, as shown by an antibody inhi bition 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 no n-integrin, RGD-independent pathway. Staurosporin-induced spreading wa s inhibited by treatment with tyrosine kinase inhibitors herbimycin A and methyl 2,5-dihydroxycinnamate. Despite the presence of staurospori n, seven proteins (220, 175, 150, 98, 62, 58, and 45 kDa) showed incre ased levels of tyrosine phosphorylation in association with cell adhes ion. Two of these (58 and 220 kDa) were identified by immunoprecipitat ion 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 no t influenced by staurosporin. Immunofluorescence microscopy revealed t hat the alpha 3 chain, diffusely expressed on the cell surface in the absence of staurosporin, was concentrated at focal adhesion plaques af ter staurosporin treatment. Neither alpha 2 nor alpha 6 was focalized by the treatment.