Figure 2.
TULA-2 dephosphorylates Syk in FcγRIIA pathway. The 50 nM anti–TULA-2 siRNA or scrambled siRNA was transiently transfected into HEL cells via nucleofection. Cells were then treated with anti-FcγRIIA clone IV.3 antibody and goat anti-mouse antibody Fab’2 (GAM) at 10 and 30 µg/mL, respectively, under stirring conditions at 37°C to crosslink and activate FcγRIIA for the indicated time. (A) At the 48-hour end point, cells were lysed, and protein was immunoblotted for TULA-2 and phosphorylated Syk at tyrosine 525/526 and 323, total Syk, FcγRIIA, and β-actin. (B-C) Phosphorylated Syk at tyrosine 525/526 and tyrosine 323 was normalized to total Syk and was plotted as mean ± standard deviation against time for the anti–TULA-2 siRNA and scrambled siRNA control groups (*P < .05, n = 5 for Y323, and n = 3 for Y525/526, 2-way analysis of variance).