N-glycoproteomics

Current research reports that lactate affects Treg metabolism, although the precise mechanism has only been partiallyelucidated. In this study, we presented evidence demonstrating that elevated lactate levels enhanced cell proliferation,suppressive capabilities, and oxidative phosphorylation (OXPHOS) in human Tregs. The expression levels of MonocarboxylateTransporters 1/2/4 (MCT1/2/4) regulate intracellular lactate concentration, thereby influencing the varying responsesobserved in naive Tregs and memory Tregs. Through mitochondrial isolation, sequencing, and analysis of human Tregs,we determined that α-1,3-Mannosyl-Glycoprotein 2-β-N-Acetylglucosaminyltransferase (MGAT1) served as the pivotaldriver initiating downstream N-glycosylation events involving progranulin (GRN) and hypoxia-upregulated 1 (HYOU1),consequently enhancing Treg OXPHOS. The mechanism by which MGAT1 was upregulated in mitochondria depended onelevated intracellular lactate that promoted the activation of XBP1s. This, in turn, supported MGAT1 transcription as well asthe interaction of lactate with the translocase of the mitochondrial outer membrane 70 (TOM70) import receptor, facilitatingMGAT1 translocation into mitochondria. Pretreatment of Tregs with lactate reduced mortality in a xenogeneic graft-versus-host disease(GvHD) model. Together, these findings underscored the active regulatory role of lactate in human Treg metabolism throughthe upregulation of MGAT1 transcription and its facilitated translocation into the mitochondria.