Modulators of inflammation use nuclear factor-KB and activator protein-1 sites to induce the caspase-1 and granzyme B inhibitor, proteinase inhibitor 9
Modulators of inflammation use nuclear factor-KB and activator protein-1 sites to induce the caspase-1 and granzyme B inhibitor, proteinase inhibitor 9
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DOI：
10.1074/jbc.m200379200
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发表时间：
2002-10-25
影响因子：
4.8
通讯作者：
Shapiro, DJ
中科院分区：
文献类型：
--
作者：
Kannan-Thulasiraman, P;Shapiro, DJ
蛋白酶抑制因子9(PI-9)抑制caspase-1(IL-1β转换酶)和颗粒酶B，从而调节促炎细胞因子IL-1β的产生和对颗粒酶B诱导的细胞凋亡的敏感性。结果表明，IL-1β、脂多糖和12-O-十四烷基佛波酯-13-乙酸酯可诱导细胞PI-9mRNA和蛋白的表达。我们在PI-9启动子区域的-135和-88处发现了功能不完善的核因子-B-K(NF-B-K)位点，在-308处发现了一致激活蛋白-1(AP-1)。利用瞬时转染法在HepG2细胞中检测PI-9启动子的突变，我们发现AP-1位点或任一NF-B-K位点的突变消融可使IL-1β诱导的PI-9的表达降低约60%。两个核因子-KB和AP-1位点的突变消融几乎取消了基础和IL-1β诱导的PI-9的表达。经IL-1β处理的HepG2细胞的核提取液显示出强烈的IL-1β诱导结合到核因子-KB和AP-1位点。凝胶迁移率分析表明，IL-1β处理后，c-Jun/c-Fos和Jund结合到AP-1位点，而p50/p65异源二聚体结合到两个NF-KB位点。雌激素诱导PI-9，但雌激素和IL-1β对PI-9的诱导不协同。在瞬时转基因的雌激素受体阳性的HepG2ER7细胞中，雌激素不干扰IL-1β的诱导，而IL-1β表现出剂量依赖性地抑制雌激素诱导的PI-9的表达。我们令人惊讶的发现是，促炎症细胞因子IL-1β强烈诱导PI-9，这表明了一种新的机制，通过控制抗炎和抗凋亡蛋白PI-9的表达来调节炎症和凋亡。
Proteinase inhibitor 9 (PI-9) inhibits caspase-1 (interleukin (IL)-1beta-converting enzyme) and granzyme B, thereby regulating production of the pro-inflammatory cytokine IL-1beta and susceptibility to granzyme B-induced apoptosis. We show that cellular PI-9 mRNA and protein are induced by IL-1beta, lipopolysaccharide, and 12-O-tetradecanoylphorbol-13-acetate. We identified functional imperfect nuclear factor-B-K (NF-B-K) sites at -135 and -88 and a consensus activator protein-1 (AP-1) site at -308 in the PI-9 promoter region. Using transient transfections in HepG2 cells to assay PI-9 promoter mutations, we find that mutational ablation of the AP-1 site or of either NF-B-K site reduces IL-1beta-induced expression of PI-9 by similar to60%. Mutational ablation of the two NF-KB sites and of the AP-1 site nearly abolishes both basal and IL-1beta-induced expression of PI-9. Nuclear extracts from IL-1beta-treated HepG2 cells exhibited strong, IL-1beta-inducible binding to the NF-KB sites and to the AP-1 site. Electrophoretic mobility shift assays show that after IL-1beta treatment c-Jun/c-Fos and JunD bind to the AP-1 site, whereas the p50/p65 heterodimer binds to the two NF-KB sites. Estrogens induce PI-9, but induction of PI-9 by estrogens and IL-1beta is not synergistic. In transiently transfected, estrogen receptor-positive HepG2ER7 cells, estrogens do not interfere with IL-1beta induction, whereas IL-1beta exhibits dose-dependent repression of estrogen-inducible PI-9 expression. Our surprising finding that the pro-inflammatory cytokine IL-1beta strongly induces PI-9 suggests a novel mechanism for regulating inflammation and apoptosis through a negative feedback loop controlling expression of the anti-inflammatory and anti-apoptotic protein, PI-9.