Figure 5.

Pretreatment with interferon-specific antibodies has differing effects on T3A-induced STAT1 phosphorylation. Cortical neuron cultures were mock or T3A infected (MOI of 100) following 4 h pretreatment with antibody directed against IFN-γ (10 μg/ml) (A). Whole-cell extracts were collected for Western blot analysis at 24 h post infection and probed for Y701-phosphorylated and total STAT1 protein levels. Recombinant IFN-γ (50 ng/ml) and T3A both induced up-regulation of total STAT1 levels and phosphorylation at Y701. Recombinant IFN-γ -induced STAT1 phosphorylation was inhibited by pretreatment with IFN-γ - directed antibody, although total STAT1 protein up-regulation remained unchanged. T3A-induced STAT1 phosphorylation and total STAT1 up-regulation was not inhibited by IFN-γ antibody pretreatment. Primary cortical neuron cultures were pretreated for 24 h with 10 μg/ml of antibody raised against IFN-γ R2 or IFN-α/βR2 before infection with T3A (MOI = 100) (B). Whole-cell extracts were prepared 24 h post infection and probed for STAT1 activation. T3A-induced STAT1 phosphorylation was unaffected by pretreatment with IFN-γ R2 antibody, similar to observations with IFN-γ antibody. Pretreatment with IFN-α/βR2 resulted in inhibition of T3A-induced STAT1 phosphorylation with no major changes in total STAT1 levels. Images provide representative observations from a minimum of four replicates per treatment.