A FUNCTIONAL SCREEN IN YEAST FOR REGULATORS AND ANTAGONIZERS OF HETEROLOGOUS PROTEIN-TYROSINE KINASES

Tyrosine phosphorylation exerts a pivotal role in cell regulation proc esses of higher eukaryotes. Tight control of the activity of protein t yrosine kinases is crucial for ordered phosphorylation to occur. We ha ve developed a functional screen for tyrosine kinase regulators using c-Src, the first cellular protein tyrosine kinase described, as a prot otype; and fission yeast, Schizosaccharomyces pombe, as a genetically amenable host system. Inducible expression of c-Src in fission yeast i s lethal. We have screened human cDNA libraries for clones able to cou nteract the lethal effect of Src. Two different classes of cDNAs, whic h we called SAS for sequences antagonizing Src, were obtained. The fir st class encodes for the protein tyrosine kinase Csk, known to regulat e Src activity through phosphorylation of the C-terminal tyrosine. The second class consists of clones encoding three different tyrosine pho sphatases, counter-acting Src action by dephosphorylation of Src subst rates and by dephosphorylation of Src itself. The system described her e can be applied to identify regulators of other heterologous tyrosine kinases, including receptor-type tyrosine kinases, which impair growt h of S. pombe.