Inhibition of ZAP70 activity impairs formation of the immunological synapse

We generated mice that express an engineered mutation of Zap-70 in the catalytic kinase domain.  This mutant “analog-sensitive” Zap-70 (or Zap-70(AS)) has kinase activity, but now is also selectively inhibited by a small molecule inhibitor called 3-MB-PP1, an analog of the kinase inhibitor PP1.  

In this figure, CD8+ Zap-70(AS) cytotoxic T lymphocytes (CTL) that formed conjugates with target cells were stained for markers of the immunological synapse.  We found that CTL were still able to form conjugates if Zap-70 was inhibited with 3-MB-PP1.  However, Zap-70 activity is required for clearance of actin from the synapse (inset) and formation of a stable immunological synapse.  As a result, inhibition of Zap-70 also severely impairs CTL cytolytic function.  Overall these studies show that Zap-70 has catalytic and non-catalytic roles in the formation of immune synapses by CTL.