Deubiquitinase USP2a Sustains Interferons Antiviral Activity by Restricting Ubiquitination of Activated STAT1 in the Nucleus
Fig 2
Ubiquitinated-pY701-STAT1 accumulates in the nucleus.
(A) 293T cells were treated with IFNα (1,000 IU/ml) for 15 min and 30 min. STAT1 ubiquitination was analyzed using the proteins from the cytoplasm and the nucleus as indicated. (B) 293T cells were treated as in (A). The proteins from the cytoplasm and the nucleus were analyzed by indicated antibodies. (C) U3A cells were transfected with Flag-STAT1-WT or Flag-STAT1-LLAA. pY701-Flag-STAT1 was immunoprecipitated using specific antibody, and immunoblotting was performed as indicated. (D) 293T cells transfected with Flag-STAT1-LLAA were stimulated with IFNα (1,000 IU/ml) for 15 min, then IFNα was removed. The proteins from the cytoplasm and the nucleus were analyzed as indicated antibodies. pY701-STAT1-Flag levels were quantificated. (E) 293T cells transfected with Flag-STAT1-WT or -LLAA were stimulated with IFNα (1,000 IU/ml) for 15 min, then IFNα was removed. The level of pY701-STAT1-Flag-WT/LLAA from whole cell lysates were analyzed and quantificated as indicated. (F) 293T cells transfected control shRNA (shCON) or KPNA1 shRNA (shKPNA1) were stimulated with IFNα (1,000 IU/ml) for 15 min, then IFNα was removed. The whole cell lysates were analyzed using indicated antibodies.