Involvement of thapsigargin- and cyclopiazonic acid-sensitive pumps in the rescue of TMEM165-associated glycosylation defects by Mn 2

FASEB J. 2019 Feb;33(2):2669-2679. doi: 10.1096/fj.201800387R. Epub 2018 Oct 11.

Abstract

Congenital disorders of glycosylation are severe inherited diseases in which aberrant protein glycosylation is a hallmark. Transmembrane protein 165 (TMEM165) is a novel Golgi transmembrane protein involved in type II congenital disorders of glycosylation. Although its biologic function is still a controversial issue, we have demonstrated that the Golgi glycosylation defect due to TMEM165 deficiency resulted from a Golgi Mn2+ homeostasis defect. The goal of this study was to delineate the cellular pathway by which extracellular Mn2+ rescues N-glycosylation in TMEM165 knockout (KO) cells. We first demonstrated that after extracellular exposure, Mn2+ uptake by HEK293 cells at the plasma membrane did not rely on endocytosis but was likely done by plasma membrane transporters. Second, we showed that the secretory pathway Ca2+-ATPase 1, also known to mediate the influx of cytosolic Mn2+ into the lumen of the Golgi apparatus, is not crucial for the Mn2+-induced rescue glycosylation of lysosomal-associated membrane protein 2 (LAMP2). In contrast, our results demonstrate the involvement of cyclopiazonic acid- and thapsigargin (Tg)-sensitive pumps in the rescue of TMEM165-associated glycosylation defects by Mn2+. Interestingly, overexpression of sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) 2b isoform in TMEM165 KO cells partially rescues the observed LAMP2 glycosylation defect. Overall, this study indicates that the rescue of Golgi N-glycosylation defects in TMEM165 KO cells by extracellular Mn2+ involves the activity of Tg and cyclopiazonic acid-sensitive pumps, probably the SERCA pumps.-Houdou, M., Lebredonchel, E., Garat, A., Duvet, S., Legrand, D., Decool, V., Klein, A., Ouzzine, M., Gasnier, B., Potelle, S., Foulquier, F. Involvement of thapsigargin- and cyclopiazonic acid-sensitive pumps in the rescue of TMEM165-associated glycosylation defects by Mn2+.

Keywords: Golgi apparatus; congenital disorders of glycosylation; manganese homeostasis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antiporters
  • Biological Transport
  • Calcium / metabolism
  • Cation Transport Proteins
  • Congenital Disorders of Glycosylation / drug therapy
  • Congenital Disorders of Glycosylation / metabolism
  • Congenital Disorders of Glycosylation / pathology
  • Endoplasmic Reticulum / drug effects
  • Endoplasmic Reticulum / metabolism
  • Enzyme Inhibitors / pharmacology
  • Gene Expression Regulation / drug effects*
  • Glycosylation
  • Golgi Apparatus / drug effects
  • Golgi Apparatus / metabolism
  • HEK293 Cells
  • Homeostasis
  • Humans
  • Indoles / pharmacology*
  • Lysosomal-Associated Membrane Protein 2 / genetics
  • Lysosomal-Associated Membrane Protein 2 / metabolism
  • Manganese / pharmacology*
  • Membrane Proteins / antagonists & inhibitors
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism*
  • Sarcoplasmic Reticulum Calcium-Transporting ATPases / genetics
  • Sarcoplasmic Reticulum Calcium-Transporting ATPases / metabolism*
  • Thapsigargin / pharmacology*

Substances

  • Antiporters
  • Cation Transport Proteins
  • Enzyme Inhibitors
  • Indoles
  • LAMP2 protein, human
  • Lysosomal-Associated Membrane Protein 2
  • Membrane Proteins
  • TMEM165 protein, human
  • Manganese
  • Thapsigargin
  • Sarcoplasmic Reticulum Calcium-Transporting ATPases
  • Calcium
  • cyclopiazonic acid