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 molecular 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. Cytostatin 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 appeared. Alkaline phosphatase treatment diminished cytostatin-induced slow-migrating paxillin. Furthermore, cytostatin increased intracellular serine/threonine-phosphorylated proteins and was found to be a selective inhibitor of protein phosphatase 2A (PP2A). Cytostatin inhibited PP2A with an IC(50) of 0.09 microgram/ml in a non-competitive manner against a substrate, p-nitrophenyl phosphate, but it had no apparent effect on other protein phosphatases including PP1, PP2B and alkaline phosphatase even at 100 microgram/ml. On the contrary, 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 modification of focal contact proteins such as paxillin by inhibiting a PP2A type protein serine/threonine phosphatase. This is the first report that describes a drug with anti-metastatic ability that inhibits PP2A selectively.
Kawada
M
M
Institute for Chemotherapy, M.C.R.F., 18-24 Miyamoto, Numazu-shi, Shizuoka, Japan.
Amemiya
M
M
Ishizuka
M
M
Takeuchi
T
T
D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology*; D002448:Cell Adhesion / Q000187:drug effects*; D003598:Cytoskeletal Proteins / Q000737:chemistry / Q000378:metabolism; D005109:Extracellular Matrix / Q000187:drug effects* / Q000378:metabolism; D008545:Melanoma; D051379:Mice; D015394:Molecular Structure; D010755:Organophosphates / Q000737:chemistry / Q000494:pharmacology*; D051419:Paxillin; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors*; D010750:Phosphoproteins / Q000737:chemistry / Q000378:metabolism; D054648:Protein Phosphatase 2; D011753:Pyrones / Q000737:chemistry / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D000903:Antibiotics, Antineoplastic; D003598:Cytoskeletal Proteins; D010755:Organophosphates; D051419:Paxillin; D010750:Phosphoproteins; C496062:Pxn protein, mouse; D011753:Pyrones; C088526:cytostatin; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2