Cytostatin, an inhibitor of cell adhesion to extracellular matrix, selectively inhibits protein phosphatase 2A

Cytostatin, which is isolated from a microbial cultured broth as a low mole cular weight inhibitor of cell adhesion to extracellular matrix (ECM), has anti-metastatic activity against B16 melanoma cells in vivo. In this study, we examined a target of cytostatin inhibiting cell adhesion to ECM. Cytost atin inhibited tyrosine phosphorylation of focal adhesion kinase (FAK) and paxillin upon B16 cell adhesion to fibronectin. While the amount of FAK was not affected by cytostatin, electrophoretically slow-migrating paxillin ap peared. Alkaline phosphatase treatment diminished cytostatin-induced slow-m igrating paxillin. Furthermore, cytostatin increased intracellular serine/t hreonine-phosphorylated proteins and was found to be a selective inhibitor of protein phosphatase 2A (PP2A). Cytostatin inhibited PP2A with an IC50 of 0.09 mu g/ml in a noncompetitive manner against a substrate, p-nitrophenyl phosphate, but it had no apparent effect on other protein phosphatases inc luding PP1, PP2B and alkaline phosphatase even at 100 mu g/ml. On the contr ary, dephosphocytostatin, a cytostatin analogue, without inhibitory effect on PP2A did not affect B16 cell adhesion including FAK and paxillin. These results indicate that cytostatin inhibits cell adhesion through modificatio n of focal contact proteins such as paxillin by inhibiting a PP2A type prot ein serine/threonine phosphatase. This is the first report that describes a drug with anti-metastatic ability that inhibits PP2A selectively. (C) 1999 Elsevier Science B.V. All rights reserved.