FIGURE 2.

Stable expression of POP2 in mouse J774A.1 macrophage regulates cytokine response and caspase-1 activation. A, flow cytometry showing intracellular staining against the myc tag in J774A.1 stable transfectants expressing Myc-pcDNA3 vector control (J7-pcDNA3) or wild-type Myc-POP2 (J7-POP2 WT). SA, secondary antibody only. B and C, J7-pcDNA3 and J7-POP2 WT cells were stimulated with indicated amounts of LPS (B) or Pam₃-CSK₄ (C) for 18 h. TNFα production was measured in culture supernatants using CBA. Cytokine results are representative of at least three experiments (error bars, S.D.; *, p < 0.05; **, p < 0.001). D, quantitative RT-PCR showing TNFα and IL-1β mRNA induction in J7-pcDNA3 and J7-POP2 WT cells in response to LPS stimulation for 1 h (TNFα) or 6 h (IL-1β). E, IL-8 expression in pcDNA3 or POP2-transfected HeLa cells in response to TNFα stimulation (10 ng/ml, 6 h). All of the mRNA results are represented as -fold induction over untreated controls and are shown as mean ± S.D. of at least three independent experiments. F, capase-1 activation was measured in J7-pcDNA3 and J7-POP2 WT cells using caspase-1 fluorescent FLICA dye and analyzed by flow cytometry. Cells were either left untreated or pulsed with 5 mm ATP for 30 min. The result shown is representative of at least three independent experiments with similar results.