SH2 domain-mediated interaction of inhibitory protein tyrosine kinase Csk with protein tyrosine phosphatase-HSCF

The protein tyrosine kinase (PTK) Csk is a potent negative regulator of sev eral signal transduction processes, as a consequence of its exquisite abili ty to inactivate Src-related PTKs, This function requires not only the kina se domain of Csk, but also its Src homology 3 (SH3) and SH2 regions. We sho wed previously that the Csk SH3 domain mediates highly specific association s with two members of the PEP family of nonreceptor protein tyrosine phosph atases (PTPs), PEP and PTP-PEST, In comparison, the Csk SH2 domain interact s with several tyrosine phosphorylated molecules, presumed to allow targett ing of Csk to sites of Src family kinase activation. Herein, we attempted t o understand better the regulation of Csk by identifying ligands for its SH 2 domain, Using a modified yeast two-hybrid screen, we uncovered the fact t hat Csk associates with PTP-HSCF, the third member of the PEP family of PTP s, This association was documented not only in yeast cells but also in a he terologous mammalian cell system and in cytokine-dependent hemopoietic cell s, Surprisingly, the Csk-PTP-HSCF interaction was found to be mediated by t he Csk SH2 domain and two putative sites of tyrosine phosphorylation in the noncatalytic portion of PTP-HSCF. Transfection experiments indicated that Csk and PTP-HSCF synergized to inhibit signal transduction by Src family ki nases and that this cooperativity was dependent on the domains mediating th eir association. Finally, we obtained evidence that PTP-HSCF inactivated Sr c-related PTKs by selectively dephosphorylating the positive regulatory tyr osine in their kinase domain. Taken together, these results demonstrate tha t part of the function of the Csk SH2 domain is to mediate an inducible ass ociation with a PTP, thereby engineering a more efficient inhibitory mechan ism for Src-related PTKs. Coupled with previously published observations, t hese data also establish that Csk forms complexes with all three known memb ers of the PEP family.