Figure 3. PAD2-mediated citrullination reciprocally regulates the activity of GATA3 and RORγt.

(A and B) Differentiated WT DBA/1J (W) and PAD2-KO (K) Th cells were restimulated with anti-CD3 (A and B) or left unstimulated (B) for 24 hours. The transcript levels of GATA3 and RORγt from 3 independent experiments are shown in A (2-tailed Student’s t test). The levels of GATA3 and RORγt proteins were analyzed with Western blotting. Representative Western blots from 5 independent experiments are shown in B. The normalized density of GATA3 and RORγt proteins is shown in the bar graphs of B (2-tailed Student’s t test). The averaged levels in WT cells without restimulation were arbitrarily set as 1. (C and D) HEK-293 cells were transfected with indicated luciferase reporters and expression vectors. Luciferase activity obtained from cells without exogenous PAD2, GATA3, and RORγt was arbitrarily set as 1 (2-tailed paired Student’s t test). Data points from the same experiments are connected with lines. Fractions of the cell extract from 2 experiments was subjected to Western blotting using indicated antibodies to demonstrate the expression of exogenous proteins and endogenous Lamin B. Representative Western blots are shown.