Metabolic Regulation of Caspases and Survival in T Cells

Caspases 的代谢调节和 T 细胞的存活

• 批准号: 9110491
• 负责人: Ralph C Budd
• 金额: $ 19.3万
• 依托单位: UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-01 至 2018-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9110491
• 关键词: Autoimmune Diseases; Autoimmune Process; CASP3 gene; Caspase; Cell Death; Cell Survival; Complex; Down-Regulation; Electron Transport; Environment; Future; Generations; Genus Hippocampus; Glycolysis; Health; Human; Immune response; Infection; Inflammatory; Interleukin-1 beta; Interleukin-15; Interleukin-2; Link; Memory; Metabolic; Metabolism; Methylation; Mitochondria; Modeling; Mouse Protein; Multiple Sclerosis; Mus; Oligomycins; Oxidative Phosphorylation; Pathogenesis; Predisposition; Protein Methylation; Proteins; Psoriasis; Reactive Nitrogen Species; Reactive Oxygen Species; Regulation; Research; Resistance; Respiration; Rheumatoid Arthritis; Role; Rotenone; Series; Site; Suggestion; Syndrome; Synovial Fluid; Synovial Membrane; T memory cell; T-Cell Proliferation; T-Lymphocyte; TNF gene; Testing; Therapeutic; Therapeutic Intervention; Time; Tissues; Ulcerative Colitis; autoreactive T cell; cytokine; driving force; extracellular; inhibitor/antagonist; mitochondrial metabolism; novel; protein expression; research study; rheumatologist; tissue culture; tool

DESCRIPTION (provided by applicant): The survival of effector T cells to the memory state is more effective for those T cells undergoing mitochondrial oxidative phosphorylation (OxPhos) rather than glycolysis. IL-15 drives OxPhos in T cells and has been associated with several autoimmune disorders, such as rheumatoid arthritis, where the inflamed synovium is rich in IL-15. It is currently unknown how IL-15 promotes OxPhos in T cells or how this promotes T cell survival. Dr. Budd's and Rincon's groups have made two key preliminary observations that offer an explanation. First, Dr. Rincon has shown that a newly defined protein known as MCJ (Methylation- Controlled J protein) negatively regulates mitochondrial Complex I activity and OxPhos (6), Second, Dr. Budd has shown that IL-15 downregulates MCJ expression, allowing increased Complex I activity, elevated OxPhos and reactive radicals compared to IL-2-cultured T cells. This results in S-nitrosylation and S- glutathionylation and inactivation of caspase-3 at critical Cys163, which resides in the enzymatic pocket. This R21 application thus posits a novel hypothesis that IL-15 promotes effector T cell survival through downregulation of MCJ, resulting in enhanced mitochondrial OxPhos, reactive radicals, and inactivation of caspase-3 through S-nitrosylation. We then apply our murine findings to an actual human inflammatory condition rich in IL-15, rheumatoid synovium. Specific Aim 1 will determine whether retroviral re-expression of MCJ in IL-15-cultured T cells inhibits mitochondrial Complex I activity and OxPhos, and increases caspase-3 activity. We will then determine whether IL-15 also promotes supercomplex formation, as observed in MCJ-/- T cells. Specific Aim 2 will define whether IL-15 present in human rheumatoid synovium promotes OxPhos, ROS/RNS, reduced caspase-3 activity and resistance to cell death through MCJ downregulation. Rheumatoid synovial tissue or synovial fluid T cells will be assessed for their metabolic state by Seahorse flux analysis, ROS/RNS generation, MCJ levels, caspase-3 activity, and resistance to cell death. This novel study combines the expertise of the PI, a research rheumatologist, in regulation of caspases by cytokines in T cells, with those of Dr. Rincon who first described MCJ in T cells and its role in regulating mitochondrial respiration at Complex I. The findings would provide the framework for future expanded studies on the role of IL-15 and MCJ in immune responses and autoimmune disorders, such as rheumatoid arthritis. All the tools are in place to conduct these studies, including MCJ-/- mice and rheumatoid synovial tissue cultures. The experiments also represent a focused exploratory study with the potential for high yield for human inflammatory conditions and suggestions for therapeutic intervention through modulation of MCJ, IL-15, and metabolism.

描述（由申请人提供）：对于那些经历线粒体氧化磷酸化（OxPhos）而不是糖酵解的T细胞，效应T细胞存活到记忆状态更有效。IL-15驱动T细胞中的OxPhos，并与几种自身免疫性疾病有关，如类风湿关节炎，其中炎症滑膜富含IL-15。目前尚不清楚IL-15如何促进T细胞中的OxPhos或如何促进T细胞存活。巴德博士和林孔的研究小组进行了两个关键的初步观察，提供了一个解释。首先，Rincon博士证明了一种新定义的蛋白质MCJ（甲基化控制J蛋白）负调控线粒体Complex I活性和OxPhos(6)。其次，Budd博士表明，与il -2培养的T细胞相比，IL-15下调MCJ表达，使Complex I活性增加，OxPhos和活性自由基升高。这导致S-亚硝基化和S-谷胱甘肽化以及位于酶袋中的关键Cys163的caspase-3失活。因此，这项R21应用提出了一个新的假设，即IL-15通过下调MCJ来促进效应T细胞的存活，从而导致线粒体OxPhos、活性自由基增强，并通过s -亚硝基化使caspase-3失活。然后，我们将我们的小鼠研究结果应用于富含IL-15的实际人类炎症状况，类风湿滑膜。特异性Aim 1将确定il -15培养的T细胞中MCJ逆转录病毒的重新表达是否抑制线粒体复合物I活性和OxPhos，并增加caspase-3活性。然后我们将确定IL-15是否也促进超复合体的形成，正如在MCJ-/- T细胞中观察到的那样。特异性Aim 2将确定IL-15是否通过MCJ下调促进人类风湿滑膜中的OxPhos、ROS/RNS、caspase-3活性降低和细胞死亡抵抗。类风湿滑膜组织或滑膜液T细胞的代谢状态将通过海马通量分析、ROS/RNS生成、MCJ水平、caspase-3活性和对细胞死亡的抗性来评估。这项新研究结合了PI（一名风湿病研究学家）在T细胞细胞因子调节半胱天冬酶方面的专业知识，以及Rincon博士的专业知识。Rincon博士首次描述了T细胞中的MCJ及其在Complex i中调节线粒体呼吸中的作用。该研究结果将为未来扩展IL-15和MCJ在免疫反应和自身免疫性疾病（如类风湿关节炎）中的作用的研究提供框架。进行这些研究的所有工具都已到位，包括MCJ-/-小鼠和类风湿滑膜组织培养。这些实验也代表了一项集中的探索性研究，具有高产量的潜力，并建议通过调节MCJ、IL-15和代谢来进行治疗干预。