A Novel Src Homology 2 Domain-containing Molecule, Src-like Adapter Protein-2 (SLAP-2), Which Negatively Regulates T Cell Receptor Signaling

Abstract

We have cloned a novel adapter protein containing Src homology 2 and Src homology 3 domains similar to the Src family of tyrosine kinases. This molecule lacks a catalytic tyrosine kinase domain and is related to a previously identified protein, Src-like adapter protein (SLAP), and is therefore designated SLAP-2. Northern blot analysis indicates that SLAP-2 is predominantly expressed in the immune system. Jurkat T cells express SLAP-2 protein and overexpression of SLAP-2 in these cells negatively regulates T cell receptor signaling as assessed by interleukin-2 promoter or NF-AT promoter reporter constructs. Mutational analysis revealed that an intact SH2 domain of SLAP-2 is essential for this inhibitory effect, whereas mutation of the SH3 domain alone has no effect. This inhibitory effect is upstream of the activation of Ras and increase of intracellular calcium levels, as no inhibition was observed when the cells were activated by phorbol ester plus ionomycin. SLAP-2 interacts with Cbl in vivo in a phosphorylation independent manner and with ZAP-70 and T cell receptor chain upon T cell receptor activation. Finally, we show that the mutation of a predicted myristoylation site within the NH 2 -terminal of SLAP-2 is essential for its inhibitory effect. This report therefore implicates SLAP and SLAP-2 as a family of adapter proteins that negatively regulate T cell receptor signaling.

Cloning and characterization of human Src-like adaptor protein 2 and a novel splice isoform, SLAP-2-v

“…While the consequences of fluctuations in the relative levels of expression of p28 and p25 SLAP-2 remain to be more fully investigated, the p25 isoform could serve to target c-Cbl to a different subcellular location or antagonize the function of p28. The p28 and p25 murine isoforms have been shown to be differentially localized to the membrane and cytoplasm, respectively, and mutation of the amino-terminal myristoylation site of p28 was shown to reduce its capacity to negatively regulate TCR-mediated NFAT activation (Holland et al, 2001;Loreto et al, 2002;Pandey et al, 2002).…”

“…The numbering refers to the coding exons. The corresponding amino acid sequences are indicated by letters above the nucleotide sequences, which start and complete each of the exons mediated NFAT activation, suggesting that SLAP-2 is a negative regulator of TCR signal transduction (Holland et al, 2001;Loreto et al, 2002;Pandey et al, 2002). Importantly, the inhibition of TCR-mediated NFAT activation by SLAP-2 is dependent on its interaction with c-Cbl (Loreto et al, 2002).…”

“…Recently, the cloning and characterization of a novel adaptor protein, Src-like adaptor protein 2 (SLAP-2), bearing significant sequence and structural similarity to SLAP, has been described (Holland et al, 2001;Loreto et al, 2002;Pandey et al, 2002). The murine SLAP-2 protein consists of an amino-terminal myristoylation sequence that is responsible for targeting SLAP-2 to cellular membranes, a Src homology 3 (SH3) domain and a Src homology 2 (SH2) domain, as well as a unique carboxy-terminal region.…”

Cutting Edge: Dexamethasone Negatively Regulates Syk in Mast Cells by Up-Regulating Src-Like Adaptor Protein

“…Relatively little is known about this protein. It is reported to negatively regulate T cell function by down-regulating expression of TCR (14) and TCR signaling by associating with Zap70 and c-Cbl (15)(16)(17)(18)(19). SLAP possesses Src homology (SH)2 and SH3 domains that are homologous to those in Src family kinases (20).…”

Normal T-Cell Development and Immune Functions in TRIM-Deficient Mice

“…For example, the TCR chain has been proposed to interact with SLAP-2 (26), TRIM (4,20), CTLA4 (7), and Unc119 (5,14), while CD3ε apparently complexes with CAST (36) and Nck (13). The physiological relevance of these interactions is so far not completely understood.…”