Figure 4.

Transfer of selected hits from the functional genetic screen to naïve Jurkat-tTA (4D9#32) cells. Diagrams of proteins predicted from the cDNA inserts and those from the corresponding wild-type genes are shown above the histograms. The left panel of histograms shows the phenotype of the original cell clones in the presence (open peaks) or absence (filled peaks) of Dox as analyzed in Figure 3a. The Dox ratio is indicated. The right top and bottom panels of histograms show the phenotypes after expressing the cDNA inserts (followed by IRES-GFP) in a naïve Jurkat-tTA population. After retroviral infection, the Jurkat-tTA (4D9#32) cells were either stimulated with the anti-TCR antibody (solid line) or left unstimulated (dashed line), and analyzed by FACS for CD69 induction after staining with anti-CD69-APC. The top right histogram in each group analyzed GFP-negative cells, which did not express the cDNA hit, whereas the bottom right histogram in each group analyzed GFP-positive cells, which expressed the cDNA hit. The following cDNA hits are shown: (a) ZAP70; (b) Lck; (c) PLC31; (d) EDG1; (e) PAK2; (f) Grb7; (g) TRAC-1.