Nck binds to the T cell antigen receptor using its SH3.1 and SH2 domains in a cooperative manner, promoting TCR functioning
20.11.2015 Paensuwan P, Hartl FA, Yousefi OS, Ngoenkam J, Wipa P, Beck-Garcia E, Dopfer EP, Khamsri B, Sanguansermsri D, Minguet S, Schamel WW, Pongcharoen S. J Immunol. 2016;196(1):448-58. In recent BIOSS publications we have shown that the proline-rich region (PRS) of the CD3epsilon subunit of the T cell antigen receptor (TCR) is important for the functioning of the TCR. Since CD3epsilon binds to the adaptor protein Nck, we cooperated with the group of Sutatip Pongcharoen, in order to analyse the biochemistry and the function of the CD3epsilon-Nck interaction. Nck contains one SH2 and three SH3 domains. Previously we have shown that the purified, recombinant SH3.1 domain of the adaptor molecule Nck can bind to the exposed PRS of CD3epsilon. Using immuno-precipitation and the in situ proximity ligation assay, we found that the binding of Nck to the TCR requires partial phosphorylation of CD3epsilon, as this interaction is based on two cooperating interactions: a) the first SH3 domain binds to the non-phosphorylated and exposed PRS, i.e. the 1st immunoreceptor tyrosine-based activation motif (ITAM) tyrosine has to be in the unphosphorylated state. B) the SH2 domain binds to the 2nd ITAM tyrosine in the phosphorylated state. Likewise, mutations of these SH3 and SH2 domains of Nck led to a loss of Nck-binding to the TCR. Furthermore, expression of an SH3-mutated Nck impaired TCR signaling. In conclusion, we demonstrate how Nck binds to the TCR and that this interaction is important for T cell activation. 