Regulation of Fas-Mediated Lung Cell Apoptosis

细胞凋亡或程序性细胞死亡调节的缺陷导致许多人类疾病，包括 肺的那些。最近的证据表明Fas（CD 95）介导的细胞凋亡在细胞凋亡中起重要作用。 几种肺部疾病的发病机制。然而，我们对所涉及的机制的理解 在这个过程中是有限的。不了解这种机制直接限制了 预防和治疗工作。本研究的总体目标是为一项 对Fas介导的细胞凋亡及其调控的分子机制的理解， 特定的肺细胞。我们的初步研究结果表明，虽然Fas可以触发肺细胞凋亡， Fas表达水平及其被Fas配体（FasL）激活与Fas易感性无关。 介导的细胞死亡，表明调节剂的凋亡信号通路必须存在。在这个项目中 我们将寻求确定控制Fas介导的肺细胞死亡的关键调节因子，并阐明其作用机制。 机制等该项目将专门测试Fas介导的细胞凋亡的易感性 和相关的肺部病理可以通过细胞FLICE抑制性的表达水平来确定。 蛋白（c-FLIP），并且这种蛋白质被某些肺毒性剂改变可以使细胞对Fas- 通过激活caspase-8和下游caspase级联介导细胞死亡。我们将确定 c-FLIP的功能作用，并使用各种方法鉴定原代肺细胞中的死亡信号通路。 分子生物学和生物化学技术。我们还将检验下调c- 通过翻译后修饰的FLIP是控制Fas介导的细胞死亡的关键调节事件 和NF-kB信号通路。因为我们的初步发现表明 活性氧和泛素-蛋白酶体依赖途径在c-FLIP降解中的作用 和Fas信号，我们将阐明潜在的机制，并确定具体的活性氧参与和他们的 细胞来源。此外，我们还将确定c-FLIP上针对该蛋白的特定泛素化位点。 使用定点诱变和基因缺失测定法对分子进行降解。预计， 拟议的研究将提供有关细胞死亡调节机制的有价值的新信息， 相关的肺部疾病，这将是重要的风险评估和治疗干预。

Defects in apoptosis or programmed cell death regulation contribute to many human diseases, including those of the lung. Recent evidence indicates that Fas(CD95)-mediated apoptosis plays an important role in the pathogenesis of several pulmonary diseases. However our understanding of the mechanisms involved in the process is limited. Failure to understand such mechanisms directly limits the effectiveness of prevention and therapeutic efforts. The overall objective of this study is to provide a scientific basis for a mechanistic understanding of the molecular events involved in Fas-mediated apoptosis and its regulation in specific lung cells. Our preliminary findings indicate that while Fas can trigger apoptosis of lung cells, the expression level of Fas and its activation by Fas ligand (FasL) do not correlate with the susceptibility toFas- mediated cell death, indicating that regulators of the apoptosis-signaling pathway must exist. In this project we will seek to identify key regulators controlling Fas-mediated cell death of lung cells and elucidate their mechanisms. The project will specifically test the hypotheses that susceptibility to Fas-mediated apoptosis and associated lung pathologies may be determined by the expression level of cellular FLICE-inhibitory protein (c-FLIP) and that alterations of this protein by certain pneumotoxic agents can sensitize cells to Fas- mediated cell death via an activation of caspase-8 and downstream caspase cascade. We will determine the functional role of c-FLIP and identify the death signaling pathways in primary lung cells using various molecular biology and biochemical techniques. We will also test the hypothesis that downregulation of c- FLIP through post-translational modifications is a critical regulatory event controlling Fas-mediated cell death and survival via caspase-8 and NF-kB signaling pathway. Because our preliminary findings indicate critical roles of reactive oxygen species (ROS) and ubiquitin-proteasome dependent pathway in c-FLIP degradation and Fas signaling, we will elucidate the underlying mechanisms and identify specific ROS involved and their cellular sources. Furthermore, we will determine specific ubiquitination sites on c-FLIP that target this molecule for degradation using site-directed mutagenesis and gene deletion assays. It is expect that the proposed studies will provide valuable new information on the mechanisms of cell death regulation and associated lung disorders which will be important in risk assessment and therapeutic intervention.