Effect of constitutive expression of SOCS3 on ISGF3 activation, STAT1 phosphorylation, and ISG43 mRNA expression by IFN-α.

SOCS3 transformants and parental KT-1/A3 cells and mock cells were incubated without (−) or with (+) 1000 U IFN-α for 30 minutes at 37°C. (A) Nuclear extracts were isolated and EMSA analysis was performed with an ISRE probe. Arrow indicates ISGF3 complex. (B) Cell extracts were prepared and immunoblotting with antiphospho-STAT1 (αPY-STAT1) or anti-STAT1 (αSTAT1) antibody was performed. Arrow indicates tyrosine phosphorylated STAT1. (C) Cells were incubated without or with 1000 U IFN-α for 1 or 4 hours. Total RNA was isolated and the expression of ISG43 was analyzed by Northern blotting.