Vanadate inhibition of protein tyrosine phosphatases in Jurkat cells: modulation by redox state

Vanadate is a potent reversible inhibitor of protein tyrosine phosphatases (PTP) in vitro. Vanadate has been shown to increase the phosphotyrosine lev els in some cell types whereas in others, like the Jurkat T-lymphoma, vanad ate has no effect. The reason for the apparent lack of effect of vanadate i n Jurkat cells was investigated in this study. Alteration of the redox stat e of these cells by reducing the glutathione level with 1-chloro-2,4-dinitr obenzene (DnpCl) had no effect on phosphotyrosine levels. However, the cell s became sensitive to vanadate, as measured by an increase in phosphotyrosi ne levels on a wide range of proteins including the MAP kinases. The increa se in phosphotyrosine levels most likely results from inhibition of cellula r PTP and suggests that protein tyrosine kinases are constitutively active in cells, resulting in a dynamic phosphorylation-dephosphorylation cycle. T he mode of inhibition of PTP by vanadate was investigated by measuring the PTP activity of Jurkat membranes isolated after treatment of cells with van adate and DnpCl. In contrast to the reversible inhibition of PTP in vitro, the effect of vanadate in the presence of DnpCl was irreversible, raising t he possibility that it is peroxovanadate formed in situ that is responsible for the inhibition of PTP in intact cells.