Niemann-Pick C1 is a late endosome-resident protein that transiently associates with lysosomes and the trans-Golgi network

Niemann-Pick type C (NPC) disease is a severe cell lipidosis characterized by the accumulation of unesterified cholesterol in the endosomal/lysosomal system. Recently the primary disease-causing gene, NPC1, was identified, bu t few clues regarding its potential function(s) could be derived from its p redicted amino acid sequence. Therefore, efforts were directed at character izing the subcellular location of the NPC1 protein. Initial studies with a FLAG-tagged NPC1 cDNA demonstrated that NPC1 is a glycoprotein that associa tes with the membranes of a population of cytoplasmic vesicles. Immunofluor escence microscopy using anti-NPC1 polyclonal antibodies confirmed this ana lysis. Double-label immunofluorescence microscopy and subcellular fractiona tion studies indicated that NPC1 associates predominantly with late endosom es (Rab9 GTPase-positive vesicles) and, to a lesser extent, with lysosomes and the trans-Golgi network. When cholesterol egress from lysosomes was blo cked by treatment of cells with U18666A, the NPC1 location shifted from lat e endosomes to the trans-Golgi network and lysosomes. Subcellular fractiona tion of liver homogenates from U18666A-treated mice confirmed these observa tions. These data suggest that U18666A may inhibit the retrograde transport of NPC1 from lysosomes to late endosomes for subsequent transfer to the tr ans Golgi network. (C) 1999 Academic Press.