Activation of Mitogen-activated Protein (MAP) Kinase Pathway by Pervanadate, a Potent Inhibitor of Tyrosine Phosphatases

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