Cation-independent mannose 6-phosphate and 78 kDa receptors for lysosomal enzyme targeting are located in different cell compartments.
The distribution of the cation-independent mannose 6-phosphate and 78 kDa receptors was studied in postnuclear subcellular fractions from two rat liver cell lines. ELISA assays revealed that the mannose 6-phosphate receptor is enriched in the light buoyant Percoll fractions that contain Golgi structures and early endosomes. Most of the 78 kDa receptor is localized in a heavy fraction at the bottom of the Percoll gradient and smaller amounts in the endosomal fractions. The high-density compartment is denser than lysosomes, contains LAMP2 but not LIMPII or acid hydrolases, and is not disrupted with glycyl-l-phenylalanine 2-naphthylamide, a substrate for cathepsin C that selectively disrupts lysosomes. Immunofluorescence microscopy studies indicate no colocalization of the 78 kDa receptor with the mannose 6-phosphate receptor or LIMPII. Mannose 6-phosphate-independent endocytosed beta-glucuronidase was found in the lysosomal, the early and late endosomal fractions. These fractions were immunoadsorbed in columns containing antibodies against the 78 kDa receptor. Only the endocytosed beta-glucuronidase present in the early and late endosomal fractions is associated to immunoadsorbed vesicles. In these vesicles, LAMP2 was detected but no LIMPII or the mannose 6-phosphate receptor. Results obtained suggest that the 78 kDa receptor is found along the endocytic pathway, but in vesicles different from the cation-independent mannose 6-phosphate receptor.
González-Noriega
Alfonso
A
Department of Cell Biology and Physiology, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, México, D.F., 04510, Mexico. gonor@biomedicas.unam.mx
Michalak
Colette
C
Antonio Sosa Melgarejo
Jorge
J
D000818:Animals; D002458:Cell Fractionation; D002460:Cell Line; D002499:Centrifugation, Density Gradient; D019084:Fluorescent Antibody Technique, Indirect; D022781:Hepatocytes / Q000378:metabolism* / Q000648:ultrastructure; D008099:Liver / Q000378:metabolism* / Q000648:ultrastructure; D008247:Lysosomes / Q000201:enzymology*; D008360:Mannosephosphates / Q000378:metabolism*; D008970:Molecular Weight; D011205:Povidone; D051381:Rats; D017527:Receptor, IGF Type 2 / Q000302:isolation & purification / Q000378:metabolism*; D012822:Silicon Dioxide; D013347:Subcellular Fractions / Q000378:metabolism*
D008360:Mannosephosphates; D017527:Receptor, IGF Type 2; C027693:mannose-6-phosphate; C016039:Percoll; D012822:Silicon Dioxide; D011205:Povidone