title: Rab proteins mediate Golgi transport of caveola-internalized glycosphingolipids and correct lipid trafficking in Niemann-Pick C cells.
abstract: We recently showed that human skin fibroblasts internalize fluorescent analogues of the glycosphingolipids lactosylceramide and globoside almost exclusively by a clathrin-independent mechanism involving caveolae. In contrast, a sphingomyelin analogue is internalized approximately equally via clathrin-dependent and caveolar routes. Here, we further characterized the caveolar pathway for glycosphingolipids, showing that Golgi targeting of sphingolipids internalized via caveolae required microtubules and phosphoinositol 3-kinases and was inhibited in cells expressing dominant-negative Rab7 and Rab9 constructs. In addition, overexpression of wild-type Rab7 or Rab9 (but not Rab11) in Niemann-Pick type C (NP-C) lipid storage disease fibroblasts resulted in correction of lipid trafficking defects, including restoration of Golgi targeting of fluorescent lactosylceramide and endogenous GM(1) ganglioside, and a dramatic reduction in intracellular cholesterol stores. Our results demonstrate a role for Rab7 and Rab9 in the Golgi targeting of glycosphingolipids and suggest a new therapeutic approach for restoring normal lipid trafficking in NP-C cells.
authors.affiliation: Department of Biochemistry and Molecular Biology, Thoracic Diseases Research Unit, Mayo Clinic and Foundation, Rochester, Minnesota 55905, USA.
authors.forename: Amit
authors.initials: A
authors.lastname: Choudhury
authors.forename: Michel
authors.initials: M
authors.lastname: Dominguez
authors.forename: Vishwajeet
authors.initials: V
authors.lastname: Puri
authors.forename: Deepak K
authors.initials: DK
authors.lastname: Sharma
authors.forename: Keishi
authors.initials: K
authors.lastname: Narita
authors.forename: Christine L
authors.initials: CL
authors.lastname: Wheatley
authors.forename: David L
authors.initials: DL
authors.lastname: Marks
authors.forename: Richard E
authors.initials: RE
authors.lastname: Pagano
mesh_terms: D015703:Antigens, CD; D001692:Biological Transport; D001896:Boron Compounds / Q000378:metabolism; D021941:Caveolae / Q000378:metabolism*; D002460:Cell Line; D002772:Cholera Toxin / Q000378:metabolism; D002784:Cholesterol / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D015870:Gene Expression; D006056:Golgi Apparatus / Q000378:metabolism*; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D007790:Lactosylceramides / Q000378:metabolism*; D050356:Lipid Metabolism; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D009542:Niemann-Pick Diseases / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D000090802:rab7 GTP-Binding Proteins
chemical_list: C095489:4,4-difluoro-4-bora-3a,4a-diaza-s-indacene; D015703:Antigens, CD; D001896:Boron Compounds; D005456:Fluorescent Dyes; D007790:Lactosylceramides; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; C413663:fluorescent protein 583; D000090802:rab7 GTP-Binding Proteins; C000716463:rab7 GTP-binding proteins, human; D049452:Green Fluorescent Proteins; C009744:CDw17 antigen; D002772:Cholera Toxin; D002784:Cholesterol; C079856:RAB9A protein, human; D020691:rab GTP-Binding Proteins