Immune Netw.  2002 Jun;2(2):72-78. 10.4110/in.2002.2.2.72.

Identification of SAP as a CTLA-4 Binding Molecule: a Role of SAP in CTLA-4 Signaling Proposed

Affiliations
  • 1Department of Pathology, University of Chicago, Chicago, IL 60637, USA. kyunglee@flowcity.bsd.uchicago.edu

Abstract

BACKGROUND: The precise mechanism by which CTLA-4 regulates T cell immune responses is still not fully understood. Previously we proposed that CTLA-4 could downregulate T cell function by modulating a signaling cascade initiated from the T cell receptor complex. The evidence for this notion comes from our findings that CTLA-4 associated with the T cell receptor zeta (TCR zeta) chain, and hence regulated TCR zeta phosphorylation by co-associated SHP-2 tyrosine phosphatase (1). In this report, we investigated whether any other signaling molecules could be involved in the CTLA-4 signaling pathway.
METHODS
We have taken biochemical approaches, such as immunoprecipitation followed by autoradiography or immunoblotting, to identify the molecules associated with CTLA-4. To perform these assays, we used activated primary T cells and ectopically transfected 293 cells. Various truncation mutants of CTLA-4 were used to map the interaction site on CTLA-4.
RESULTS
We found that in addition to TCR zeta and SHP-2, a recently cloned small adaptor molecule, SAP (SLAM-associated protein), was also able to associate with CTLA-4. We identified the domain of SAP association in CTLA-4 being a motif involving GVYVKM. This motif has been previously found to bind SHP-2 through its phosphorylated tyrosine interaction with SH-2 domain of SHP-2. Indeed, co-expression of SAP and SHP-2 reduced their binding to CTLA-4 significantly, suggesting that SAP and SHP-2 compete for the common binding site, GVYVKM. Thus, by blocking SHP-2 recruitment SAP could function as a negative regulator of CTLA-4.
CONCLUSION
Taken together, our data suggest the existence of complicate signaling cascade in regulating CTLA-4 function, and further provide evidence that SAP can act either as a positive or negative regulator depending on the nature of the associating receptors.

Keyword

CTLA-4; SAP; SHP-2; tyrosine kinase; transfection; T cells

MeSH Terms

Autoradiography
Binding Sites
Clone Cells
Immunoblotting
Immunoprecipitation
Phosphorylation
Protein Tyrosine Phosphatase, Non-Receptor Type 11
Protein-Tyrosine Kinases
Receptors, Antigen, T-Cell
T-Lymphocytes
Transfection
Tyrosine
Protein Tyrosine Phosphatase, Non-Receptor Type 11
Protein-Tyrosine Kinases
Receptors, Antigen, T-Cell
Tyrosine
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