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）：效应T细胞与记忆状态的存活更为有效，对于那些接受线粒体氧化磷酸化（OXPHOS）而不是糖酵解的T细胞。 IL-15驱动T细胞中的Oxphos，并与几种自身免疫性疾病（例如类风湿关节炎）相关，其中发炎的滑膜在IL-15中富含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。 Budd's和Rincon博士的团体进行了两个关键的初步观察，提供了解释。首先，Rincon博士表明，新定义的称为MCJ（甲基化控制的J蛋白）负调节线粒体复合物I活性和Oxphos（6），其次，Budd博士表明IL-15降低MCJ表达，使IL-15的表达下调，允许IL-15的表达，使IL-15的IL-15下调，使IL的复杂I活性增加，氧气和反应性自由基与IL-2-Cult-cult-2-cult-culted t t t t t t te confectials iveption i活性。这会导致cys163处的caspase-3的S-亚硝基化和谷胱甘肽化和灭活，该caspase-3位于酶袋中。因此，这种R21的应用提出了一个新的假设，即IL-15通过下调MCJ促进效应T细胞的存活，从而导致线粒体oxphos，反应性自由基和通过S-硝基化的Caspase-3失活。然后，我们将鼠的发现应用于富含IL-15的类风湿滑膜的实际人类炎症状况。具体目标1将确定在IL-15培养的T细胞中MCJ的逆转录病毒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是否还促进了MCJ - / - T细胞中观察到的超复合形成。特定的目标2将定义人类类风湿滑膜中存在的IL-15是否促进OXPHOS，ROS/RN，降低CASPASE-3活性以及通过MCJ下调对细胞死亡的耐药性。将通过海马通量分析，ROS/RNS产生，MCJ水平，caspase-3活性以及对细胞死亡的耐药性评估类风湿滑膜组织或滑液T细胞的代谢状态。 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关节炎。所有工具都可以进行这些研究，包括MCJ - / - 小鼠和类风湿滑膜组织培养物。该实验还代表了一项集中的探索性研究，具有通过调节MCJ，IL-15和代谢的调节，具有高收率的人类炎症条件和治疗干预的建议。