MicroRNA-27b Contributes to Lipopolysaccharide-mediated Peroxisome Proliferator-activated Receptor γ (PPARγ) mRNA Destabilization
MicroRNA-27b Contributes to Lipopolysaccharide-mediated Peroxisome Proliferator-activated Receptor γ (PPARγ) mRNA Destabilization
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DOI：
10.1074/jbc.m109.066399
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发表时间：
2010-04-16
影响因子：
4.8
通讯作者：
Bruene, Bernhard
中科院分区：
文献类型：
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作者：
Jennewein, Carla;von Knethen, Andreas;Bruene, Bernhard
过氧化物酶体增殖物激活受体γ（PPAR γ）作为慢性炎症性疾病的治疗靶点获得了相当大的兴趣。值得注意的是，诸如多发性硬化症或阿尔茨海默病等疾病的发病机制与受损的PPAR γ表达相关。考虑到炎症过程中PPAR γ表达的调节在很大程度上是未知的，我们有兴趣阐明潜在的机制。为此，我们通过将原代人巨噬细胞暴露于脂多糖（LPS）来启动炎症反应，并观察到PPAR γ 1表达的快速下降。由于启动子活性不受LPS的影响，我们专注于mRNA的稳定性，并注意到mRNA的半衰期缩短。由于RNA稳定性通常通过3 '-非翻译区（UTR）调节，因此我们使用特定构建体通过报告基因测定分析了PPAR γ-3'-UTR的影响。LPS显著降低了pGL 3-PPARgamma-3 '-UTR的荧光素酶活性，表明PPARgamma 1 mRNA是不稳定的。3 '-UTR内潜在的微小RNA-27 a/B（miR-27 a/B）结合位点的缺失或突变恢复了荧光素酶活性。此外，在LPS暴露后诱导的miR-27 b的抑制部分逆转了PPAR γ 1 mRNA的衰减，而miR-27 b过表达降低了PPAR γ 1 mRNA的含量。此外，LPS进一步减少了这种衰减。miR-27 b的抑制或过表达证实了miR-27 b依赖性PPAR γ 1减少的功能相关性，miR-27 b的抑制或过表达影响了LPS诱导的促炎细胞因子肿瘤坏死因子α（TNF α）和白细胞介素（IL）-6的表达。我们提供的证据表明，LPS诱导的miR-27 b有助于不稳定的PPAR γ 1 mRNA。了解降低PPAR γ的分子机制可能有助于更好地了解炎症性疾病。
Peroxisome proliferator-activated receptor gamma (PPAR gamma) gained considerable interest as a therapeutic target during chronic inflammatory diseases. Remarkably, the pathogenesis of diseases such as multiple sclerosis or Alzheimer is associated with impaired PPAR gamma expression. Considering that regulation of PPAR gamma expression during inflammation is largely unknown, we were interested in elucidating underlying mechanisms. To this end, we initiated an inflammatory response by exposing primary human macrophages to lipopolysaccharide (LPS) and observed a rapid decline of PPAR gamma 1 expression. Because promoter activities were not affected by LPS, we focused on mRNA stability and noticed a decreased mRNA half-life. As RNA stability is often regulated via 3'-untranslated regions (UTRs), we analyzed the impact of the PPAR gamma-3'-UTR by reporter assays using specific constructs. LPS significantly reduced luciferase activity of the pGL3-PPAR gamma-3'-UTR, suggesting that PPAR gamma 1 mRNA is destabilized. Deletion or mutation of a potential microRNA-27a/b (miR-27a/b) binding site within the 3'-UTR restored luciferase activity. Moreover, inhibition of miR-27b, which was induced upon LPS exposure, partially reversed PPAR gamma 1 mRNA decay, whereas miR-27b overexpression decreased PPAR gamma 1 mRNA content. In addition, LPS further reduced this decay. The functional relevance of miR-27b-dependent PPAR gamma 1 decrease was proven by inhibition or overexpression of miR-27b, which affected LPS-induced expression of the pro-inflammatory cytokines tumor necrosis factor alpha (TNF alpha) and interleukin (IL)-6. We provide evidence that LPS-induced miR-27b contributes to destabilization of PPAR gamma 1 mRNA. Understanding molecular mechanisms decreasing PPAR gamma might help to better appreciate inflammatory diseases.