Dephosphorylation of ZAP-70 and inhibition of T cell activation by activated SHP1

Abstract

Studies with motheaten mice, which lack the SHP1 protein tyrosine phosphatase, indicate that this enzyme plays an important negative role in T cell antigen receptor (TCR) signaling. The physiological substrates for SHP1 in T lymphocytes, however, have remained unclear or controversial. To define these targets for SHP1 we have compared the effects of constitutively active and inactive mutants of SHP1 on TCR signaling. Expression of wild‐type SHP1 had a very small effect on the TCR‐induced tyrosine phosphorylation of ZAP‐70 and Syk, even when SHP1 was overexpressed 20 – 100‐fold over endogenous SHP1. Inactive SHP1‐D421A and wild‐type SHP2 were without effects. Constitutively active SHP1‐ΔSH2 had a more pronounced effect on ZAP‐70 and Syk, even when expressed at near physiological levels. SHP1‐ΔSH2 also inhibited events downstream of ZAP‐70 and Syk, such as activation of the mitogen‐activated protein kinase Erk2 and the transcriptional activation of the interleukin‐2 gene. In contrast, a constitutively active SHP2‐ΔSH2 had no statistically significant effect (although it caused a slight augmentation in some individual experiments). None of the constructs influenced the anti‐CD3‐induced tyrosine phosphorylation of the TCR ζ‐chain or phospholipase Cγ1, indicating that Src family kinase function was intact. Taken together, our findings support the notion that ZAP‐70 and Syk can be direct substrates for SHP1 in intact cells. However, the two SH2 domains of SHP1 did not facilitate its recognition of ZAP‐70 and Syk as substrates in intact cells. Therefore, we suggest that SHP1 is not actively recruited to inhibit TCR signaling induced by ligation of this receptor alone. Instead, we propose that ligation of a distinct inhibitory receptor leads to the recruitment of SHP1 via its SH2 domains, activation of SHP1 and subsequently inhibition of TCR signals if the inhibitory receptor is juxtaposed to the TCR.

The Cell-Cell Adhesion Molecule Carcinoembryonic Antigen-Related Cellular Adhesion Molecule 1 Inhibits IL-2 Production and Proliferation in Human T Cells by Association with Src Homology Protein-1 and Down-Regulates IL-2 Receptor

“…In this respect, we have shown by cocapping experiments that CEACAM1 does indeed associate with the TCR complex, and that small amounts of ZAP-70 are coimmunoprecipitated with CEACAM1 from activated PBMCs. These results agree with studies by Mustelin and coworkers (33), who showed that SHP-1, but not SHP-2, can dephosphorylate ZAP-70 and inhibit T cell activation. In other studies, these investigators showed that SHP1 remains in the cytosol and requires activation for its inhibitory activity (32).…”

“…Low levels of ZAP-70 were found associated with CEACAM1 before and after ligation with T84.1 with a 2-fold increase after ligation. The association of CEACAM1 with both SHP-1 and ZAP-70 is of special significance given that SHP-1-deficient mice have hyperactive TCR responses (30,31) and ZAP-70 has been shown to be a target of SHP-1 (32,33). The results for CaM were even more dramatic, with a 5-fold increase of binding after CEACAM1 ligation.…”

Treatment of Human T Cells with Bisperoxovanadium Phosphotyrosyl Phosphatase Inhibitors Leads to Activation of Cyclooxygenase-2 Gene

“…The fact that bpV-mediated activation of COX-2 promoter was still detected in the CD45-negative cell line J45.01 is an indication that this T cell membrane PTP is not participating to this process (data not shown). The Src homology-containing PTP SHP-1 represents an ideal candidate, because it has been proposed as a negative regulator of T cell activation by decreasing the activity of several molecules involved in TCR signaling machinery, including p56 lck , phospholipase C␥, ZAP-70, and Syk (25,30,(53)(54)(55). Our data showing an inhibition of bpVinduced transcriptional activation when SHP-1 is overexpressed further strengthen this hypothesis.…”

TCR-induced downregulation of protein tyrosine phosphatase PEST augments secondary T cell responses

“…Human LYP, PTPH1, SHP1, LMPTP-B and HePTP cDNA in mammalian expression vector pEF5HA were described previously (Bottini et al, 2002;Brockdorff et al, 1999;Gjörloff-Wingren et al, 2000;Saxena et al, 1998;Vang et al, 2005). The HD-PTP cDNA was a gift from Dr. M. Ouchida (Okayama University, Japan; Toyooka et al, 2000) …”