Phosphorylation of STAT1 and STAT2 by members of the JAK tyrosine kinase family is a prerequisite for activation of these transcription factors to drive type I IFN-induced gene expression. Therefore, we analyzed whether STAT phosphorylation patterns are altered in influenza A virus infected cells that were stimulated with IFN at different time points post infection (p.i.). The human alveolar epithelial cell line A549 was infected with the influenza A virus strain A/Puerto-Rico/8/34 (H1N1) (PR8) ( Figure 1A ). Cells were subsequently stimulated with IFNb at given time-points p.i. and STAT phosphorylation was assessed in Western blots. Both STAT1 and STAT2 were readily phosphorylated upon cytokine stimulation in uninfected cells or in infected cells up to 4 h p.i. ( Figure 1A ). Furthermore, virus infection alone resulted in a significant induction of STAT phosphorylation 4-6 h p.i., presumably caused by virus-induced IFN expression. However, at later time points (6-10 h p.i.), in A549 cells both virus-and IFN-induced STAT1 and STAT2 phosphorylation was markedly reduced ( Figure 1A ). Similar patterns were observed upon stimulation of cells with IFNa or upon infection with other viruses, such as the human influenza virus A/Victora/3/75 (H3N2) (data not shown). In addition, this phenomenon could also be detected in other epithelial cells such as the human embryonic kidney cell line HEK293 ( Figure 2E ) or the human umbilical vein endothelial cells (HUVEC) ( Figure S1B ). Inhibition was not caused by indirect disturbing effects on cellular metabolism or enzyme activities due to ongoing virus replication, since IFNc-induced STAT1 phosphorylation was not affected at all ( Figure 1C ). Finally, involvement of any auto-or paracrine action of virus-induced type I IFN could be ruled out, as the inhibitory effect was also observed in Vero cells lacking functional type I IFN genes ( Figure 1E ).