Noncleavable poly(ADP-ribose) polymerase-1 regulates the inflammation response in mice
J. Clin. Invest. Virginie Pétrilli, et al. 114:1072 doi:10.1172/JCI21854 [Go to this article.]

Figure 5
Noncleavable PARP-1 impairs NF-κB–dependent gene expression in response to proinflammatory stimuli. (A) EMSA analysis of NF-κB binding activity in PARP-1^+/+ and PARP-1^KI/KI macrophage extracts. This experiment was repeated at least 5 times using 3 independent sets of samples. (B) Primary PARP-1^+/+ MLFs and PARP-1^KI/KI MLFs were transfected with the indicated luciferase reporter vectors: nos-2(1485/+31WT)-Luc (nos-2 WT-Luc) and nos-2(1485/+31NF-κBmut)-Luc (nos-2 mutκB-Luc). The luciferase activity was measured after stimulation by LPS and/or IFN-γ for 12 hours. (C) Primary PARP-1^–/– peritoneal macrophages were cotransfected with nos-2(1485/+31WT)-Luc or nos-2(1485/+31NF-κBmut)-Luc, together with vectors expressing wild-type PARP-1 (WT) or caspase-resistant PARP-1 (D₂₁₄N; upper panel) or enzyme-dead PARP-1 (M890V/D899N; lower panel). NF-κB activation (fold increase) in B and C was determined by the ratio of the luciferase activity between cells treated and untreated with LPS and IFN-γ. The value of untreated cells was arbitrarily set to 1. Error bars indicate standard errors of 3 independent experiments. (D) Western blot analysis shows corresponding PARP-1 proteins used in panel C. (E) Western blot analysis of PARP-1 cleavage in THP-1 cells upon LPS stimulation. Note that PARP-1 cleavage products are readily visible 6 hours after LPS stimulation. This Western blot is representative of the 2 experiments.