Figure 9

MAPKs activation by L. pneumophila through flagellin and inhibition of L. pneumophila -induced CREB and ATF1 activation and IL-8 transcription by p38 inhibitor. (A) Jurkat cells were infected with Corby or flaA mutant (MOI, 100:1), and lysates were subjected to immunoblotting. Cells were pretreated with the indicated concentrations of SB203580 for 1 h prior to infection and subsequently infected with Corby (MOI, 100:1) for 4 h (B) and for 6, 8, 12, or 24 h (C). IL-8 mRNA expression on the harvested cells was analyzed by RT-PCR (B) and the supernatants were subjected to ELISA to determine IL-8 secretion (C). (D) Cells were transfected with -133-luc and then pretreated with the indicated concentrations of SB203580 for 1 h prior to infection. They were infected subsequently with Corby for 6 h. Luciferase (LUC) activity was assayed. The solid bar indicates LUC activity of -133-luc without infection. (E) Cells were transfected with -133-luc and dominant negative mutants and then infected with Corby for 6 h. The solid bar indicates LUC activity of -133-luc without infection. All values were calculated as the change (n-fold) in induction values relative to the basal level measured in uninfected cells. Data are mean ± SD of three experiments. (F) Cells were pretreated with or without SB203580 (50 μM) for 1 h prior to infection and subsequently were infected with Corby (MOI, 100:1). Lysates were subjected to immunoblotting. dn, dominant negative.