MTHFD2 ablation in T cells protects against heart transplant rejection by perturbing IRF4/PD-1 pathway through the metabolic-epigenetic nexus

One-carbon metabolism supports the activation, proliferation, and function of multiple immune cells. However, researchers have not clearly determined whether and how one-carbon metabolic enzymes contribute to heart transplant rejection.We investigated the dynamic metabolic adaptation in grafts during heart transplant rejection by conducting transcriptomics, metabolomics and single-cell RNA sequencing studies of cardiac tissue from human and mouse heart transplant recipients. We also assessed the expression of the one-carbon metabolic enzyme methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) in cardiac grafts by immunofluorescence and flow cytometry assays. Then we constructed a murine heart transplant model with T cell-specific Mthfd2 knockout mice, analyzed T cells function by flow cytometry assays and enzyme-linked immunospot assays, and studied the mechanism by Cleavage Under Targets and Tagmentation assays. Finally, we studied the effect of a pharmacological inhibitor of MTHFD2 in humanized skin transplant model.We revealed that the one-carbon metabolism enzyme MTHFD2 was a hallmark of alloreactive T cells and was linked to T cell proliferation and function after exposure to alloantigen. And, Mthfd2 ablation prevented murine heart transplant rejection. Mechanistically, we found Mthfd2 ablation affected the interferon regulatory factor 4/programmed death-1 pathway through a metabolic-epigenetic mechanism involving H3K4me3. Furthermore, we found that inhibiting MTHFD2 attenuated human allograft rejection in a humanized skin transplant model.These data show that the one-carbon metabolic enzyme MTHFD2 serves as a metabolic checkpoint of alloreactive T cells and suggest that it may be a potential therapeutic target for heart transplant rejection.