The Role of Intracellular Serpins in the Regulation of Necrosis

描述(申请人提供)：坏死是许多疾病的共同病理特征，但关于这一途径执行的分子细节在很大程度上仍不清楚。我们的实验室对坏死性细胞死亡的机制有了重要的见解，检测了细胞内丝氨酸蛋白的抗蛋白酶功能。利用线虫模型系统，已经证明SRP-6在响应多种应激诱导刺激时具有阻断坏死的功能。最近的研究表明，缺乏SRP-6会导致蛋白质动态平衡受损，而蛋白毒性的增加会导致野生型或SRP-6缺失动物的坏死性细胞死亡。这个项目的长期目标是剖析蛋白毒性和坏死性细胞死亡之间的关系，并确定参与这些过程调控的特定基因。线虫模型系统将被用来研究以下具体目标：1)确定SRP-6的丢失如何扰乱蛋白质稳态；2)确定扰乱蛋白质稳态如何导致坏死性细胞死亡。在目标1中，我将在SRP-6缺失的动物中进行救援实验，以确定SRP-6抑制活性是否参与蛋白质稳态的调节。此外，我将使用RNAi筛选来确定特定的多肽酶作为SRP-6的调控靶点。在目标2中，我将利用正向和反向遗传方法来识别调节蛋白质稳态和/或NCD途径的特定基因。这些研究的结果将为调节蛋白质稳态和坏死的机制提供新的见解，有助于开发治疗坏死相关疾病的治疗方案。 公共卫生相关性：坏死是许多疾病的共同病理特征，包括与缺血性损伤、慢性炎症和许多胃肠道疾病相关的疾病。这项研究计划的目标是确定导致坏死性细胞死亡的分子机制和识别关键基因。最终，这些研究将为开发针对坏死相关疾病患者的高度特异性治疗方法提供基础。

DESCRIPTION (provided by applicant): Necrosis is a common pathologic feature of numerous diseases, but the molecular details regarding the execution of this pathway remain largely unknown. Our lab has produced significant insight into the mechanisms involved in necrotic cell death examining the anti-protease function of intracellular serpin proteins. Using the C. elegans model system, it has been shown that SRP-6 functions to block necrosis in response to multiple stress-inducing stimuli. More recent studies demonstrate that lack of SRP-6 results in impaired protein homeostasis, and that increased proteotoxicity results in necrotic cell death in either wild- type or srp-6 null animals. The long term objectives of this project are to dissect the relationship between proteotoxicity and necrotic cell death, and to identify specific genes involved in the regulation of these processes. The C. elegans model system will be used to examine the following specific aims: 1) to determine how the loss of SRP-6 perturbs protein homeostasis and 2) to determine how perturbed protein homeostasis results in necrotic cell death. In Aim 1 I will perform rescue experiments in srp-6 null animals to determine whether SRP-6 inhibitory activity is involved in the regulation of protein homeostasis. Additionally, I will employ an RNAi screen to identify specific peptidases as SRP-6 regulatory targets. In Aim 2, I will utilize both forward and reverse genetic approaches to identify specific genes that modulate the protein homeostasis and/or NCD pathways. The results of these studies will provide new insights into the mechanisms that regulate protein homeostasis and necrosis, facilitating the development of therapeutic options for treatment of necrosis-related diseases. PUBLIC HEALTH RELEVANCE: Necrosis is a common pathologic feature of many diseases including those associated with ischemic injury, chronic inflammation, and many gastrointestinal disorders. The goal of this research proposal is to determine the molecular mechanisms and to identify critical genes that contribute to necrotic cell death. Ultimately, these studies will provide the basis for development of highly specific treatments for patients with necrosis-associated disorders.