Efficient T-cell receptor signaling requires a high-affinity interaction between the Gads C-SH3 domain and the SLP-76 RxxK motif

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Abstract
The relationship between the binding affinity and specificity of modular interaction domains is potentially important in determining biological signaling responses. In signaling from the T‐cell receptor (TCR), the Gads C‐terminal SH3 domain binds a core RxxK sequence motif in the SLP‐76 scaffold. We show that residues surrounding this motif are largely optimized for binding the Gads C‐SH3 domain resulting in a high‐affinity interaction ( K D=8–20 nM) that is essential for efficient TCR signaling in Jurkat T cells, since Gads‐mediated signaling declines with decreasing affinity. Furthermore, the SLP‐76 RxxK motif has evolved a very high specificity for the Gads C‐SH3 domain. However, TCR signaling in Jurkat cells is tolerant of potential SLP‐76 crossreactivity, provided that very high‐affinity binding to the Gads C‐SH3 domain is maintained. These data provide a quantitative argument that the affinity of the Gads C‐SH3 domain for SLP‐76 is physiologically important and suggest that the integrity of TCR signaling in vivo is sustained both by strong selection of SLP‐76 for the Gads C‐SH3 domain and by a capacity to buffer intrinsic crossreactivity.