Mutant Rab7 causes the accumulation of cathepsin D and cation-independent mannose 6-phosphate receptor in an early endocytic compartment.
Stable BHK cell lines inducibly expressing wild-type or dominant negative mutant forms of the rab7 GTPase were isolated and used to analyze the role of a rab7-regulated pathway in lysosome biogenesis. Expression of mutant rab7N125I protein induced a dramatic redistribution of cation-independent mannose 6-phosphate receptor (CI-MPR) from its normal perinuclear localization to large peripheral endosomes. Under these circumstances approximately 50% of the total receptor and several lysosomal hydrolases cofractionated with light membranes containing early endosome and Golgi markers. Late endosomes and lysosomes were contained exclusively in well-separated, denser gradient fractions. Newly synthesized CI-MPR and cathepsin D were shown to traverse through an early endocytic compartment, and functional rab7 was crucial for delivery to later compartments. This observation was evidenced by the fact that 2 h after synthesis, both markers were more prevalent in fractions containing light membranes. In addition, both were sensitive to HRP-DAB- mediated cross-linking of early endosomal proteins, and the late endosomal processing of cathepsin D was impaired. Using similar criteria, the lysosomal membrane glycoprotein 120 was not found accumulated in an early endocytic compartment. The data are indicative of a post-Golgi divergence in the routes followed by different lysosome-directed molecules.
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Department of Biochemistry, Molecular Biology and Cell Biology, Northwestern University, Evanston, Illinois 60208-3500, USA.
Feng
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Y
Hoflack
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Wandinger-Ness
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D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D002402:Cathepsin D / Q000096:biosynthesis / Q000378:metabolism*; D002412:Cations; D002458:Cell Fractionation; D002460:Cell Line; D002462:Cell Membrane; D006224:Cricetinae; D004705:Endocytosis / Q000502:physiology*; D011992:Endosomes / Q000201:enzymology; D019204:GTP-Binding Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D008024:Ligands; D051907:Lysosomal Membrane Proteins; D008361:Mannosidases / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism; D016296:Mutagenesis; D017527:Receptor, IGF Type 2 / Q000378:metabolism*; D014162:Transfection; D001619:beta-N-Acetylhexosaminidases / Q000378:metabolism; D020691:rab GTP-Binding Proteins; D000090802:rab7 GTP-Binding Proteins
D015703:Antigens, CD; D002412:Cations; D008024:Ligands; D051907:Lysosomal Membrane Proteins; D008562:Membrane Glycoproteins; D017527:Receptor, IGF Type 2; D000090802:rab7 GTP-Binding Proteins; D008361:Mannosidases; C038829:mannosyl-oligosaccharide 1,3 - 1,6-alpha-mannosidase; D001619:beta-N-Acetylhexosaminidases; D002402:Cathepsin D; D019204:GTP-Binding Proteins; D020691:rab GTP-Binding Proteins