The laboratory has spent a significant time investigating cytokine regulation of protein phosphorylation. All the cytokines investigated to date induce the activation of both serine and tyrosine kinase activity. Comparative analysis of the substrates phosphorylated by the proliferative cytokines revealed that consensus substrates are phosphorylated in response to these individual ligands suggesting that they utilize certain components common to the signal transduction processes (with the respective receptor systems). A 68 kDa protein is phosphorylated on serine residues in response to proliferative cytokines and was identified as L-plastin. IL 2 was also shown to regulate the serine phosphorylation of the anti-oncogene retinoblastoma gene product RB, as well as to activate the cell cycle serine kinase p34CDC2. During the course of these studies we have also discovered a previously unknown serine kinase which is associated with cell cycle arrest. The 55 kDa serine kinase has functional expression that may qualify it as a new class of anti-oncogene. All the cytokines tested also induced tyrosine phosphorylation of a number of proteins. We found that among the cytokines tested a number of phospho- tyrosyl substrates were identical by Mr, isoelectric mobility and peptide mapping. The laboratory has focused its attention to one of these substrates p97, which was phosphorylated on tyrosine by stimulating cells with IL 2, IL 3, GM-CSF, EPO, or prolactin. We have examined three receptor complexes for the presence of tyrosine kinase activity and phosphotyrosyl substrates. Using biotinylated ligands or antibody towards the IL 2R, EPO-R, and prolactin receptor, we found tyrosine kinase activity associated within these receptor complexes. Our efforts to characterize the nature of the catalytic protein within these receptor complexes have suggested that the p97 protein has properties consistent with tyrosine kinases. We are pursuing the purification and molecular cloning of this enzyme in order to determine its functional role in IL 2R and EPO-R initiated signal transduction.
NIH ExPorter
NIH ExPorter