title: Endomannosidase undergoes phosphorylation in the Golgi apparatus.
abstract: Glucose residues from N-linked oligosaccharides are removed by glucosidases I and II in the endoplasmic reticulum (ER) or by the alternate endomannosidase pathway in the Golgi apparatus. Our morphological analysis demonstrates that recombinant rat endomannosidase exhibited a cis- and medial-Golgi localization alike the endogenous enzyme and its ER to Golgi transport is COP II mediated. Recombinant endomannosidase undergoes a posttranslational modification, which is not related to N-or O-glycosylation. A shift in molecular mass of recombinant endomannosidase was observed upon phosphatase digestion but not for ER-retained CHO cell endomannosidase. Furthermore, immunoprecipitation of (35)S- and (33)P-labeled endomannosidase expressed in CHO-K1 cells suggests that recombinant endomannosidase undergoes phosphorylation. Substitution of the single cytoplasmic threonine residue of rat endomannosidase by either an alanine or valine residue resulted in the same posttranslational modification alike the wild-type enzyme. The subcellular localization and the in vivo activity of the mutant endomannosidase were not affected. Thus, endomannosidase phosphorylation is occurring in luminal sequences. Modification was prevented when endomannosidase was synthesized using reticulocyte lysates in the presence of canine microsomes. Treatment of cells with brefeldin A blocked the posttranslational modification of endomannosidase, suggesting that phosphorylation is occurring in the Golgi apparatus, the residence of endomannosidase.
authors.affiliation: Division of Cell and Molecular Pathology, Department of Pathology, University of Zurich, CH-8091 Zurich, Switzerland.
authors.forename: Tania
authors.initials: T
authors.lastname: Torossi
authors.forename: Bruno
authors.initials: B
authors.lastname: Guhl
authors.forename: Jürgen
authors.initials: J
authors.lastname: Roth
authors.forename: Martin
authors.initials: M
authors.lastname: Ziak
mesh_terms: D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D005786:Gene Expression Regulation; D006031:Glycosylation; D006056:Golgi Apparatus / Q000737:chemistry / Q000378:metabolism*; D007150:Immunohistochemistry; D037102:Lectins / Q000737:chemistry; D008361:Mannosidases / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry*; D016253:Microscopy, Immunoelectron / Q000379:methods; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D051381:Rats; D000520:alpha-Glucosidases / Q000737:chemistry
chemical_list: D037102:Lectins; D008565:Membrane Proteins; C034251:4-nitrophenyl-alpha-glucosidase; D008361:Mannosidases; C529770:endomannosidase, rat; D000520:alpha-Glucosidases