JCS02822 Supplementary Material

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• Supplemental Figure 1 -

  Fig. S1. Constitutive nuclear import of STAT2 is independent of tyrosine phosphorylation. (A) LMB treatment did not induce STAT2-GFP tyrosine phosphorylation. NIH3T3 cells stably expressing STAT2-GFP were stimulated for 2 hours with IFN-b, LMB, or left untreated. Whole-cell extracts were subjected to immunoprecipitation with anti-STAT2 antibody, resolved by SDS-PAGE, transferred to a membrane and probed with phosphotyrosine specific antibody PY-20. The membrane was stripped and reprobed with anti-(Tyr701)Phospho STAT1 antibody followed by antibody against the STAT2 C-terminal. (B) LMB-induced nuclear accumulation of STAT2 was not abolished when the activating kinases were blocked. NIH3T3 cells stably expressing STAT2-EGFP were pretreated with staurosporine (500 nM) for 1 hour. LMB plus staurosporine (a) or staurosporine alone (b) was then added and fluorescence imaging of the cells was done 1 hour later. (C) LMB-induced nuclear accumulation of the activation-deficient mutant STAT2R601K-GFP in NIH3T3 cells.