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The Vav binding site (Y315) in ZAP-70 is critical for antigen receptor-mediated signal transduction
J Wu1, Q Zhao, T Kurosaki
1Department of Microbiology and Immunology, University of California, San Francisco, California 94143, USA.
Abstract:
Stimulation of antigen receptors in T and B cells leads to the activation of the Src and Syk families of protein tyrosine kinases (PTK). These PTKs subsequently phosphorylate numerous intracellular substrates, including the 95-kD protooncogene product Vav. Vav is essential for both T and B cell development and T and B cell antigen receptor-mediated signal transduction. After receptor ligation, Vav associates with phosphorylated Syk and ZAP-70 PTKs, an interaction that depends upon its SH2 domain. Here we demonstrate that a point mutation of tyrosine 315 (Y315F) in ZAP-70, a putative Vav SH2 domain binding site, eliminated the Vav- ZAP-70 interaction. Moreover, the Y315 mutation impaired the function of ZAP-70 in antigen receptor signaling. Surprisingly, this mutation also resulted in marked reduction in the tyrosine phosphorylation of ZAP-70, Vav, SLP-76, and Shc. These data demonstrate that the Vav binding site in ZAP-70 plays a critical role in antigen receptor-mediated signal transduction.
Insights
The Vav binding site on ZAP-70 is crucial for T and B cell antigen receptor signaling. Mutation of this site disrupts ZAP-70 function and downstream signaling pathways.
Area of Science:
- Immunology
- Molecular Biology
- Cell Signaling
Background:
- T and B cell activation relies on protein tyrosine kinases (PTKs) like Src and Syk.
- Vav, a key substrate, is vital for T and B cell development and signaling.
- Vav interacts with phosphorylated Syk and ZAP-70 PTKs via its SH2 domain.
Purpose of the Study:
- To investigate the role of the Vav SH2 domain binding site on ZAP-70 in antigen receptor signaling.
- To elucidate the functional consequences of disrupting the Vav-ZAP-70 interaction.
Main Methods:
- Site-directed mutagenesis was used to create a Y315F mutation in ZAP-70.
- The impact of this mutation on Vav-ZAP-70 interaction was assessed.
- Antigen receptor signaling pathways were analyzed in cells with the mutated ZAP-70.
Main Results:
- A Y315F mutation in ZAP-70 abolished the interaction with Vav.
- The Y315F mutation impaired ZAP-70 function in antigen receptor signaling.
- This mutation significantly reduced the tyrosine phosphorylation of ZAP-70, Vav, SLP-76, and Shc.
Conclusions:
- The Vav binding site on ZAP-70 is critical for effective antigen receptor-mediated signal transduction.
- Disruption of this interaction impacts multiple signaling components, highlighting its central role.