Abstract
The intraperitoneal injection of a vanadate/H2O2 mixture (peroxovanadate) into mice resulted within minutes in the appearance of numerous tyrosine-phosphorylated proteins in the liver and kidney. These effects are presumably due to the inhibition of phosphotyrosine phosphatase activity. Three of the tyrosine-phosphorylated proteins have been identified as the receptors for epidermal growth factor, insulin, and hepatocyte growth factor. The injection of peroxovanadate also enhanced the tyrosine phosphorylation of many of the proteins known to function downstream of these receptors, including SHC, signal transducer and activator of transcription (Stat) 1alpha,beta, Stat 3, Stat 5, phospholipase C-gamma, insulin receptor substrate 1, GTPase-activating protein, beta-catenin, gamma-catenin, p120cas, SHP-1, and SHP-2. The administration of peroxovanadate also induced nuclear translocation of a number of tyrosine-phosphorylated Stat proteins. In addition, the global effects on tyrosine phosphorylation permitted the detection of a number of novel intracellular protein interactions, including an association of Tyk2 with beta-catenin. The in situ administration of peroxovanadate may prove useful in the search for novel tyrosine-phosphorylated proteins and the identification of new interactions between previously identified tyrosine-phosphorylated substrates.
Highlights
Many of the cellular responses induced by hormones, growth factors, and cytokines are mediated by the activation of intracellular kinases
We have previously reported that the intraperitoneal injection of EGF into mice leads to a rapid increase in the tyrosine phosphorylation of a number of proteins in all organs examined [16]
In this report we demonstrate that the intraperitoneal injection of vanadate/H2O2 mixtures into mice, in the absence of any added ligand, induces the rapid and massive tyrosine phosphorylation of multiple cellular proteins in both the liver and kidney
Summary
Many of the cellular responses induced by hormones, growth factors, and cytokines are mediated by the activation of intracellular kinases. To detect tyrosine-phosphorylated proteins in cell extracts, specific phosphatase inhibitors, such as sodium vanadate [1], must be added.
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