ACTIVATION OF MITOGEN-ACTIVATED PROTEIN (MAP) KINASE PATHWAY BY PERVANADATE, A POTENT INHIBITOR OF TYROSINE PHOSPHATASES

Rapid tyrosine phosphorylation of key cellular pro teins is a crucial event in signal transduction. The regulatory role of protein-tyrosine phosphatases (PTPs) in this process was explored by studying the effec ts of a powerful PTP inhibitor, pervanadate, on the activation of the mitogen-activated protein (MAP) kinase cascade. Treatment of HeLa cell s with pervanadate resulted in a marked inhibition of PTP activity, ac companied by a drastic increase in tyrosine phosphorylation of cellula r proteins. The increased tyrosine phosphorylation coincided with the activation of the MAP kinase cascade as indicated by enzymatic activit y assays of MEK (MAP kinase/ERK-kinase) and MAP kinase and gel mobilit y shift analyses of Raf-1 and MAP kinase. The activation was sustained but reversible. Upon removal of pervanadate, both tyrosine phosphoryl ation and MAP kinase activation declined to basal levels. Therefore, i nhibition of PTP activity is sufficient per se to initiate a complete MAP kinase activation program.