Tat inhibits the nuclear translocation of IFNγ-activated pSTAT1 and the expression of ISGs. (A) PBMos were untreated (UT), mock treated (M), or treated with indicated concentrations of Tat (T) for 4 hours and followed by the treatment of IFNγ (100 U/mL) for 5 minutes. Cell lysates were examined by Western blotting using anti-pJak1 or -pJak2 antibodies. Total Janus kinases were probed as loading control. The levels of phosphorylation were normalized with total Jak1 and Jak2, respectively, whereas the fold changes of individual treatment with reference to the untreated were indicated. (B) PBMos were pretreated with mock (M) or Tat (T) for 4 hours and then stimulated with IFNγ (100 U/mL) as indicated on top of each panel. Nuclear extracts harvested were examined by Western blotting using anti-pSTAT1 antibodies. Lamin B1 was probed as loading control. (C) PBMos were untreated (UT), mock treated (M), or Tat treated (T, 10 nM) for 4 hours and followed by the treatment of IFNγ (100 U/mL) for 16 hours. The expression of IFNγ-inducible genes was examined using RT-PCR. GAPDH was used as a loading control.