Introduction: Current research reports that lactate affects Treg metabolism, although the precise mechanism remains partially understood. In this study, we present evidence demonstrating that elevated lactate levels significantly enhance cell proliferation, suppressive capabilities, and oxidative phosphorylation (OXPHOS) in human Tregs. The expression levels of Monocarboxylate Transporters 1/2/4 (MCT1/2/4) regulate intracellular lactate concentration, thereby influencing the varying responses observed in naive Tregs and memory Tregs. Through mitochondrial isolation, sequencing, and analysis of human Tregs, we have determined that N-acetylglucosaminyltransferase I(MGAT1) serves as the pivotal driver initiating downstream N-glycosylation events involving GRN and HYOU1, consequently enhancing Treg OXPHOS. The mechanism by which MGAT1 is upregulated in mitochondria depends on two key factors: Elevated intracellular lactate promotes the activation of XBP1s, which, in turn, supports MGAT1 transcription. Furthermore, lactate interacts with the translocase of the mitochondrial outer membrane 70 (TOM70) import receptor, facilitating the translocation of MGAT1 into the mitochondria. Pre-treatment of Tregs with lactate significantly reduces mortality in a xenogeneic graft-versus-host disease (GvHD) model. These findings underscore the active regulatory role of lactate in human Treg metabolism through the upregulation of MGAT1 transcription and its facilitated translocation into the mitochondria.

Method: Total proteomics, protein glycosylation modification omics, lentiviral transfection, luciferase assay, et al. 

Results: Lactate promotes suppressive function and OXPHOS of memory and naïve Tregs. A high lactate environment increases MGAT1 expression in memory and naïve Tregs by mitochondrial proteomics. Lactate increases N-glycosylation of GRN and HYOU1 to promote Treg OXPHOS via MGAT1. Mitochondrial function of Treg declines after MGAT1 knockdown. A high lactate environment elevates XBP1s facilitates MGAT1 transcription. TOM70 is essential for the mitochondrial translocation of MGAT1 in high lactate environment. MGAT1 knockdown attenuates the therapeutic effect of Treg in a Xeno-GvHD model.