title: Acidification of the Golgi apparatus is indispensable for maturation but not for cell surface delivery of Ret.
abstract: We examined the effect of concanamycin A and bafilomycin A1, inhibitors of the vacuolar proton-ATPase, on maturation and expression of Ret, a tyrosine kinase receptor for glial cell line-derived neurotrophic factor. Ret appeared as 150- and 170-kDa bands on sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels and both forms were sensitive to peptide-N-glycosidase F. Western and immunocytochemical analyses revealed that the 150-kDa immature form of Ret accumulated in the Golgi apparatus upon treatment with vacuolar proton-ATPase inhibitors, whereas, the 170-kDa mature form of Ret was dramatically decreased. The result suggests that glycosylation of Ret during the conversion from immature forms to mature forms is pH sensitive, and is likely initiated in the acidic trans-Golgi apparatus. In contrast, glycosylation of nascent receptors to become immature receptors appeared to be pH insensitive, and are likely to take place in the endoplasmic reticulum. The immature form of Ret was present in the plasma membrane when the cells were treated with the vacuolar proton-ATPase inhibitors. In conclusion, the acidification of the Golgi apparatus is crucial for maturation of Ret but not indispensable for trafficking of receptors to the membrane.
authors.affiliation: Department of Biomolecular Science, Faculty of Engineering, Gifu University, Yanagido, Gifu, Japan. yokoh@gifu-u.ac.jp
authors.forename: Yoko
authors.initials: Y
authors.lastname: Hirata
authors.forename: Norihiro
authors.initials: N
authors.lastname: Shimokawa
authors.forename: Kentaro
authors.initials: K
authors.lastname: Oh-hashi
authors.forename: Zu-Xi
authors.initials: ZX
authors.lastname: Yu
authors.forename: Kazutoshi
authors.initials: K
authors.lastname: Kiuchi
mesh_terms: D000818:Animals; D019921:Biotinylation; D015153:Blotting, Western; D002462:Cell Membrane / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D066246:ErbB Receptors / Q000378:metabolism; D006056:Golgi Apparatus / Q000502:physiology*; D007150:Immunohistochemistry; D018942:Macrolides / Q000494:pharmacology; D016716:PC12 Cells; D051096:Proto-Oncogene Proteins c-ret / Q000378:metabolism*; D051381:Rats; D020917:Receptor, trkA / Q000096:biosynthesis / Q000235:genetics; D011956:Receptors, Cell Surface / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D018942:Macrolides; D011956:Receptors, Cell Surface; C036978:concanamycin A; C040929:bafilomycin A1; D066246:ErbB Receptors; D051096:Proto-Oncogene Proteins c-ret; D020917:Receptor, trkA
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