Nck recruitment to the TCR required for ZAP70 activation during thymic development.?

et al.

The adaptor protein Nck is inducibly recruited through its SH3.1 domain to a proline-rich sequence (PRS) in CD3 after TCR engagement. However, experiments with a knockin mutant bearing an 8-aa replacement of the PRS have indicated that Nck binding to the TCR is constitutive, and that it promotes the degradation of the TCR in preselection double-positive (DP) CD4+ CD8+ thymocytes. To clarify these discrepancies, we have generated a new knockin mouse line (KI-PRS) bearing a conservative mutation in the PRS resulting from the replacement of the two central prolines. Thymocytes of KI-PRS mice are partly arrested at each step at which pre-TCR or TCR signaling is required. The mutation prevents the trigger-dependent inducible recruitment of endogenous Nck to the TCR but does not impair TCR degradation. However, KI-PRS preselection DP thymocytes show impaired tyrosine phosphorylation of CD3z, as well as impaired recruitment of ZAP70 to the TCR and impaired ZAP70 activation. Our results indicate that Nck is recruited to the TCR in an inducible manner in DP thymocytes, and that this recruitment is required for the activation of early TCR-dependent events. Differences in the extent of PRS mutation could explain the phenotypic differences in both knockin mice. Copyright © 2013 by The American Association of Immunologists, Inc.

Comision Interministerial de Ciencia y Tecnologia (Grant SAF2010-14912); Redes Temáticas de Investigación Cooperativa Sanitaria (Grant RD06/0020/1002); Fundación Científica Asociación Española Contra el Cáncer; European Union (Grant FP7/2007-2013 “SYBILLA”); Deutsche Forschungsgemeinschaft (Grant SFB620); Bundesministerium fur Bildung; Wissenschaft, Forschung und Technologie (Grant 01 EO 0803); Fundación Ramón Areces.