N -Glycosylation of MRS2 balances aerobic and anaerobic energy production by reducing rapid mitochondrial Mg 2+ influx in conditions of high glucose or impaired respiratory chain function.

Autor: Peng M; Mitochondrial Medicine Frontier Program, Division of Human Genetics, Department of Pediatrics, The Children's Hospital of Philadelphia, Philadelphia, PA 19104., Mathew ND; Mitochondrial Medicine Frontier Program, Division of Human Genetics, Department of Pediatrics, The Children's Hospital of Philadelphia, Philadelphia, PA 19104., Anderson VE; Mitochondrial Medicine Frontier Program, Division of Human Genetics, Department of Pediatrics, The Children's Hospital of Philadelphia, Philadelphia, PA 19104., Falk MJ; Mitochondrial Medicine Frontier Program, Division of Human Genetics, Department of Pediatrics, The Children's Hospital of Philadelphia, Philadelphia, PA 19104.; Department of Pediatrics, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104., Nakamaru-Ogiso E; Mitochondrial Medicine Frontier Program, Division of Human Genetics, Department of Pediatrics, The Children's Hospital of Philadelphia, Philadelphia, PA 19104.; Department of Pediatrics, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104.
Jazyk: angličtina
Zdroj: BioRxiv : the preprint server for biology [bioRxiv] 2024 Jul 09. Date of Electronic Publication: 2024 Jul 09.
DOI: 10.1101/2024.07.09.602756
Abstrakt: N -linked glycoproteins function in numerous biological processes, modulating enzyme activities as well as protein folding, stability, oligomerization, and trafficking. While N -glycosylation of mitochondrial proteins has been detected by untargeted MS-analyses, the physiological existence and roles of mitochondrial protein N -linked glycosylation remain under debate. Here, we report that MRS2, a mitochondrial inner membrane protein that functions as the high flux magnesium transporter, is N -glycosylated to various extents depending on cellular bioenergetic status. Both N -glycosylated and unglycosylated isoforms were consistently detected in mitochondria isolated from mouse liver, rat and mouse liver fibroblast cells (BRL 3A and AFT024, respectively) as well as human skin fibroblast cells. Immunoblotting of MRS2 showed it was bound to, and required stringent elution conditions to remove from, lectin affinity columns with covalently bound concanavalin A or Lens culinaris agglutinin. Following peptide: N -glycosidase F (PNGase F) digestion of the stringently eluted proteins, the higher M r MRS2 bands gel-shifted to lower M r and loss of lectin affinity was seen. BRL 3A cells treated with two different N -linked glycosylation inhibitors, tunicamycin or 6-diazo-5-oxo-l-norleucine, resulted in decreased intensity or loss of the higher M r MRS2 isoform. To investigate the possible functional role of MRS2 N - glycosylation, we measured rapid Mg 2+ influx capacity in intact mitochondria isolated from BRL 3A cells in control media or following treatment with tunicamycin or 6-diazo-5-oxo-l-norleucine. Interestingly, rapid Mg 2+ influx capacity increased in mitochondria isolated from BRL 3A cells treated with either N -glycosylation inhibitor. Forcing reliance on mitochondrial respiration by treatment with either galactose media or the glycolytic inhibitor 2-deoxyglucose or by minimizing glucose concentration similarly reduced the N -glycosylated isoform of MRS2, with a correlated concomitant increase in rapid Mg 2+ influx capacity. Conversely, inhibiting mitochondrial energy production in BRL 3A cells with either rotenone or oligomycin resulted in an increased fraction of N -glycosylated MRS2, with decreased rapid Mg 2+ influx capacity. Collectively, these data provide strong evidence that MRS2 N -glycosylation is directly involved in the regulation of mitochondrial matrix Mg 2+ , dynamically communicating relative cellular nutrient status and bioenergetic capacity by serving as a physiologic brake on the influx of mitochondrial matrix Mg 2+ under conditions of glucose excess or mitochondrial bioenergetic impairment.
Competing Interests: M L., D.I., C.R., P.K., C.B., N.D.M., R.X., C.S., E.N.O. and V.E.A., have no relevant financial disclosures. M.J.F. is an inventor on US Patent No. PCT/US 17/256,406 entitled, “Compositions and Methods for Treatment of Mitochondrial Respiratory Chain Dysfunction and Other Mitochondrial Disorders,” filed in the Name of The Children’s Hospital of Philadelphia on 12/28/20. M.J.F is co-founder and chief scientific advisor of Rarefy Therapeutics, a scientific advisory board member with equity interest in RiboNova, Inc., and scientific board member as paid consultant with Khondrion and with Larimar Therapeutics. M.J.F. has previously been or is currently engaged as a paid consultant with Abliva [formerly Neurovive], Astellas [formerly Mitobridge] Pharma Inc., Casma Therapeutics, Cyclerion Therapeutics, Epirium Bio, HealthCap VIII Advisor AB, Imel Therapeutics, Minovia Therapeutics, Reneo Therapeutics, Stealth BioTherapeutics, Taysha Therapeutics, Zogenix, Inc. and/or as a sponsored research collaborator with AADI Therapeutics, Astellas [formerly Mitobridge] Pharma Inc., Cyclerion Therapeutics, Epirium Bio [formerly Cardero Therapeutics], Imel Therapeutics, Merck, Minovia Therapeutics Inc., Mission Therapeutics, NeuroVive, Raptor Therapeutics, REATA Inc., Reneo Therapeutics, RiboNova Inc., Standigm Therapeutics, and Stealth BioTherapeutics. M.J.F has received royalties from Elsevier and a speaker’s honorarium from PlatformQ and Agios Pharma.
Databáze: MEDLINE
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