IDENTIFICATION OF P130(CAS) AS A SUBSTRATE FOR THE CYTOSOLIC PROTEIN-TYROSINE-PHOSPHATASE PTP-PEST

PTP-PEST is a ubiquitously expressed, cytosolic, mammalian protein tyr osine phosphatase (PTP) which exhibits high specific activity in vitro , We have investigated the substrate specificity of PTP-PEST by a nove l substrate-trapping approach in combination within vitro dephosphoryl ation experiments. We initially identified a prominent 130-kDa tyrosin e-phosphorylated protein in pervanadate-treated HeLa cell lysates whic h was preferentially dephosphorylated by PTP-PEST in vitro, In order t o identify this potential substrate, mutant (substrate-trapping) forms of PTP-PEST were generated which lack catalytic activity but retain t he ability to bind substrates. These mutant proteins associated in sta ble complex-es exclusively with the same 130-kDa protein, which was id entified as p130(cas) by immunoblotting. This exclusive association wa s observed in lysates from several cell lines and in transfected COS c ells, but was not observed with other members of the PTP family, stron gly suggesting that p130(cas) represents a major physiologically relev ant substrate for PTP-PEST. Our studies suggest potential roles for PT P-PEST in regulation of p130(cas) function, These functions include mi togen- and cell adhesion-induced signalling events and probable roles in transformation by various oncogenes. These results provide the firs t demonstration of a PTP having an inherently restricted substrate a s pecificity in vitro and in vivo. The methods used to identify p130(cas ) as a specific substrate for PTP-PEST are potentially applicable to a ny PTP and should therefore prove useful in determining the physiologi cal substrates of other members of the PTP family.