Adaptative response to inflammatory stressors

早期的研究表明，转录因子STAT 3参与了对白细胞介素-6家族细胞因子和生长因子的急性期应答负责的靶基因的诱导表达。以前已经证明，STAT 3蛋白是翻译后修饰的靶标，例如磷酸化、谷胱甘肽化和乙酰化，所有这些都增加了细胞外信号对STAT 3调节的层。例如，我们报道了STAT 3在用小硫醇化合物吡咯烷二硫代氨基甲酸酯（PDTC）处理细胞后对S-谷胱甘肽化敏感，从而阻止了IL-6介导的STAT 3磷酸化、二聚化和与DNA的结合。此外，STAT 3通过多种酶在胞质溶胶和细胞核中发生可逆乙酰化，使得响应于细胞因子刺激，组蛋白乙酰转移酶p300/CREB结合蛋白与STAT 3缔合并促进其乙酰化。相反，包括SIRT 1在内的几种组蛋白脱乙酰酶与STAT 3相互作用，并通过关键STAT 3赖氨酸残基的脱乙酰化来负调控其转录活性。目前尚不清楚STAT 3的其他细胞功能是否涉及SIRT 1。 在这项研究中，Sirt 1基因敲除小鼠胚胎成纤维细胞（MEF）细胞被用来描绘的作用，SIRT 1的表达和亚细胞定位的STAT 3和线粒体功能的各种标志物。我们证明，Sirt 1-null细胞有显着更高的丝氨酸磷酸化STAT 3水平的线粒体，与线粒体生物能量学和ATP的形成增加。各种线粒体生物能量参数进行了测量，如在细胞培养物中的耗氧率，在分离的线粒体中的电子传递链复合物的酶活性，和生产的ATP和乳酸。两种独立的方法，包括SIRT 1的异位表达和siRNA介导的STAT 3敲低，导致Sirt 1-KO细胞中细胞内ATP减少和乳酸产生增加，达到接近野生型对照的水平。我们还进行了广泛的磷酸化抗体阵列分析，并进一步建立了Sirt 1-null细胞中促炎NF- B通路的组成性激活。重要的是，通过p65 Rel基因敲低抑制NF- B活性阻断了STAT 3基因诱导，同时降低了Sirt 1-KO细胞中的细胞呼吸。这些结果表明，NF- B的激活可能在通过线粒体STAT 3的更高表达和激活触发Sirt 1缺失细胞中增加细胞呼吸的促炎程序中起重要作用。这些结果对于理解STAT 3和SIRT 1在促炎性疾病中的相互作用具有重要意义。 转录因子HSF 1通过应激反应机制控制寿命，使适应性反应的物理和炎症的压力。我们最近报道PDTC部分通过HSF 1介导的转录激活赋予细胞抗炎症保护作用。HSF 1是否有助于SIRT 1的表达和/或激活，并通过非基因组STAT 3功能改变线粒体满足能量需求的能力仍有待确定。这项工作可能会导致对抗慢性炎症性疾病和延长寿命的新方法。

Earlier work has demonstrated that the transcription factor, STAT3, is involved in the inducible expression of target genes responsible for acute-phase response in response to the interleukin-6 family of cytokines and growth factors. It has previously demonstrated that the STAT3 protein is a target for post-translational modifications such as phosphorylation, glutathionylation and acetylation, all of which adding layers of STAT3 regulation by extracellular signals. For example, we reported that STAT3 is susceptible to S-glutathionylation after cell treatment with the small thiol compound, pyrrolidine dithiocarbamate (PDTC), thus preventing the IL-6-mediated phosphorylation, dimerization and binding of STAT3 to DNA. Moreover, STAT3 is subject to reversible acetylation that occurs both in the cytosol and the nucleus by multiple enzymes, such that in response to cytokine stimulation, the histone acetyltransferase p300/CREB-binding protein associates with STAT3 and promotes its acetylation. Conversely, several histone deacetylases, including SIRT1, interact with STAT3 and negatively regulate its transcriptional activity through deacetylation of key STAT3 lysine residues. It is unclear whether other cellular functions of STAT3 involve SIRT1. In this study, Sirt1 gene knockout murine embryonic fibroblasts (MEF) cells were used to delineate the role of SIRT1 in the expression and subcellular localization of STAT3 and various markers of mitochondrial function. We demonstrate that Sirt1-null cells have significantly higher serine phosphorylated STAT3 level in mitochondria that correlated with increase in mitochondrial bioenergetics and formation of ATP. Various mitochondrial bioenergetic parameters were measured, such as the oxygen consumption rate in cell cultures, enzyme activities of the electron transport chain complexes in isolated mitochondria, and production of ATP and lactate. Two independent approaches, including ectopic expression of SIRT1 and siRNA-mediated knockdown of STAT3, led to reduction in intracellular ATP and increased lactate production in Sirt1-KO cells to levels that were approaching those of wild-type controls. We also performed extensive phospho-antibody array analysis and further establish a constitutive activation of the pro-inflammatory NF-κB pathway in Sirt1-null cells. Of significance, suppression of NF-κB activity by p65Rel gene knockdown blocked STAT3 gene induction while reducing cellular respiration in Sirt1-KO cells. These results suggest that the activation of NF-κB may play an important role in triggering a pro-inflammatory program of increased cellular respiration in Sirt1-null cells via higher expression and activation of mitochondrial STAT3. These results have implications for understanding the interplay between STAT3 and SIRT1 in pro-inflammatory conditions. The transcription factor HSF1 controls longevity through a stress-response mechanism, enabling adaptive responses to physical and inflammatory stressors. We recently reported that PDTC confers cellular protection against inflammation partly via HSF1-mediated transcriptional activation. Whether HSF1 contributes to the expression and/or activation of SIRT1 and alters the capacity of mitochondria to meet energy demands through non-genomic STAT3 functions remains to be established. This work may lead to new ways of fighting chronic inflammatory diseases and increase longevity.