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.
The survival of effector T cells to the memory state is of fundamental importance to protection from infection as well as in autoimmune disorders. IL-15 has been implicated in the pathogenesis of various autoimmune diseases such as rheumatoid arthritis, multiple sclerosis, ulcerative colitis, celiac syndrome, and psoriasis. IL- 15 also indues the inflammatory cytokines TNF-? and IL-1? and is important in homeostatic proliferation of T cells. It is now appreciated that augmented homeostatic proliferation may be a driving force in several autoimmune disorders. IL-15 also drives a metabolic state characterized by high oxidative phosphorylation and low glycolysis, and we observe that this is associated with low levels of caspase-3 activity due to its inactivation by S-nitrosylation. Regulation of active caspases in cycling T cells is paramount to understanding T cell survival during both homeostatic proliferation as well as during the generation of memory T cells. We have also identified a novel protein, MCJ as a regulator of Complex I activity in the electron transport chain, and hence of mitochondrial respiration. IL-15 silences MCJ expression. The significance of the proposed studies is that it links for the first time the cytokine environment to metabolism via MCJ, and regulation of effector caspase-3 as a major determinant of cell survival. It then applies this to human rheumatoid arthritis synovium. The study presents a focused series of experiments on the metabolic regulation of caspase-3 activity by IL-15 and applies this to a real human autoimmune disease. It also suggests potential therapeutic options in autoimmune disorders by regulating IL-15, MCJ, or metabolism.
|Secinaro, Michael A; Fortner, Karen A; Dienz, Oliver et al. (2018) Glycolysis promotes caspase-3 activation in lipid rafts in T cells. Cell Death Dis 9:62|
|Barbier-Torres, Lucía; Iruzubieta, Paula; Fernández-Ramos, David et al. (2017) The mitochondrial negative regulator MCJ is a therapeutic target for acetaminophen-induced liver injury. Nat Commun 8:2068|