Triple arginines in the cytoplasmic tail of endomannosidase are not essential for type II membrane topology and Golgi localization.
Endomannosidase is a Golgi-localized endoglycosidase, which provides an alternate glucosidase-independent pathway of glucose trimming. Using a protease protection assay we demonstrated that Golgi-endomannosidase is a type II membrane protein. The first 25 amino acids of this protein, containing the cytoplasmic tail and the transmembrane domain, were sufficient for Golgi retention of fused reporter proteins alpha1-antitrypsin or green fluorescent protein. However, shortening or deletion of the transmembrane domain prevented Golgi localization, while lengthening it partially reduced Golgi retention of the enzyme. Substitution of the highly conserved positively charged amino acids within the cytoplasmic tail had neither an effect on type II topology nor on the inherent Golgi localization of the enzyme. In contrast, cytoplasmic tail-deleted rat endomannosidase possessed an inverted topology resulting in endoplasmic reticulum mislocalization. Thus, proper topology rather than the presence of positively charged amino acids in the cytoplasmic tail is critical for Golgi localization of rat endomannosidase.
Stehli
J
J
Division of Cell and Molecular Pathology, Department of Pathology, University of Zurich, Schmelzbergstrasse 12, Zurich, Switzerland.
Torossi
T
T
Ziak
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine / Q000502:physiology*; D016466:CHO Cells; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D006056:Golgi Apparatus / Q000378:metabolism / Q000648:ultrastructure*; D006367:HeLa Cells; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism / Q000648:ultrastructure*; D008361:Mannosidases / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology; D008954:Models, Biological; D021382:Protein Sorting Signals / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology; D021381:Protein Transport; D051381:Rats
D008565:Membrane Proteins; D021382:Protein Sorting Signals; D001120:Arginine; D008361:Mannosidases; C529770:endomannosidase, rat