Magnesium transporter 1 (MAGT1) deficiency causes selective defects in N-linked glycosylation and expression of immune-response genes

Abstract

Magnesium transporter 1 (MAGT1) critically mediates magnesium homeostasis in eukaryotes and is highly-conserved across different evolutionary branches. In humans, loss-offunction mutations in the MAGT1 gene cause X-linked magnesium deficiency with Epstein-Barr virus (EBV) infection and neoplasia (XMEN), a disease that has a broad range of clinical and immunological consequences. We have previously shown that EBV susceptibility in XMEN is associated with defective expression of the antiviral natural-killer group 2 member D (NKG2D) protein and abnormal Mg 2؉ transport. New evidence suggests that MAGT1 is the human homolog of the yeast OST3/ OST6 proteins that form an integral part of the N-linked glycosylation complex, although the exact contributions of these perturbations in the glycosylation pathway to disease pathogenesis are still unknown. Using MS-based glycoproteomics, along with CRISPR/Cas9-KO cell lines, natural killer cell-killing assays, and RNA-Seq experiments, we now demonstrate that humans lacking functional MAGT1 have a selective deficiency in both immune and nonimmune glycoproteins, and we identified several critical glycosylation defects in important immune-response proteins and in the expression of genes involved in immunity, particularly CD28. We show that MAGT1 function is partly interchangeable with that of the paralog protein tumorsuppressor candidate 3 (TUSC3) but that each protein has a different tissue distribution in humans. We observed that MAGT1dependent glycosylation is sensitive to Mg 2؉ levels and that reduced Mg 2؉ impairs immune-cell function via the loss of specific glycoproteins. Our findings reveal that defects in protein glycosylation and gene expression underlie immune defects in an inherited disease due to MAGT1 deficiency.MAGT1 is an evolutionally conserved Mg 2ϩ -specific ion transport facilitator found in all animals and has been shown to participate in the multienzyme complex responsible for enzymatic coupling of N-glycans onto peptide substrates (1, 2). Null mutations in the MAGT1 gene lead to the rare primary immunodeficiency "X-linked immunodeficiency with Mg 2ϩ defect, Epstein-Barr virus (EBV) 7 infection and neoplasia" (XMEN) disease (3,4). Here, we explore these dual roles by examining cells from both healthy and MAGT1-deficient humans.Mg 2ϩ is the most abundant divalent cation in eukaryotic cells, with intracellular concentrations ranging from 15 to 20 mM depending on the cell type. Most Mg 2ϩ is tightly bound to cellular substituents, especially nucleic acids, nucleoside triphosphates, and enzymes. The unbound intracellular free Mg 2ϩ is estimated to be 0.4 -1.0 mM or ϳ1-5% of the total Mg 2ϩ concentration in the cell (5, 6), and because Mg 2ϩ is the biologically active form of Mg, these intracellular concentra-

Defective glycosylation and multisystem abnormalities characterize the primary immunodeficiency XMEN disease

“…Interestingly, we found defective glycosylation of certain ion transporters in XMEN samples, and Mg 2+ deficiency can affect glycosylation (14). The MAGT1 gene is conserved in unicellular eukaryotes such as yeast, indicating an ancient phylogenetic connection between glycosylation and Mg 2+ (12).…”

“…Protein glycosylation is a posttranslational modification critical for normal immune function (11). MAGT1 has high amino acid sequence homology with the human tumor suppressor candidate 3 protein (TUSC3) and the yeast oligosaccharyl transferase 3/6 (OST3/6) proteins that participate in the enzymatic complex that performs asparagine N-linked glycosylation (NLG) in the endoplasmic reticulum (ER) (12)(13)(14). Each OST complex has 1 catalytic subunit, either STT3A or STT3B, and multiple noncatalytic subunits creating distinct but complementary NLG enzyme complexes (15,16).…”

“…Conversely, STT3B favors sequons that would be difficult to glycosylate cotranslationally, including those in the terminal 50-55 amino acids of the carboxyl tail and short loops between TM regions (17,18). MAGT1 can associate with the STT3Bcontaining OST complex and promote NLG of STT3B-dependent glycoproteins in human tumor cell lines (14,19). Genetic diseases affecting protein glycosylation, congenital disorders of glycosylation (CDG), can involve genes that add glycans to proteins in the ER (type I) or further process protein-bound glycans in the Golgi apparatus (type II) (20,21).…”

A Review of the Action of Magnesium on Several Processes Involved in the Modulation of Hematopoiesis

“…As the molecular relationship to OST subunits and complexities of Mg 2+ suggested that pivotal manifestations of XMEN syndrome may rotate around defective glycosylation, in addition to or instead of alterations in Mg 2+ transport [ 171 ]. However, a study from the same group also showed that glycosylation is sensitive to Mg 2+ levels and that reduced Mg 2+ impairs immune cell function via the loss of specific glycoproteins [ 172 ].…”

Magnesium, Oxidative Stress, Inflammation, and Cardiovascular Disease

“…MagT1 deficiency leads to the absence of a T cell antigen receptor-stimulated Mg flux and attenuation of T cell activation [ 155 , 173 ]. Matsuda-Lennikov et al have reported that MagT1 mediates N-linked glycosylation and expression of immune-response genes in T cells [ 174 ].…”