Biosynthesis, glycosylation, movement through the Golgi system, and transport to lysosomes by an N-linked carbohydrate-independent mechanism of three lysosomal integral membrane proteins.
The biosynthesis, glycosylation, movement through the Golgi system, transport to lysosomes, and turnover of three lysosomal integral membrane proteins (LIMPSs) have been studied in normal rat kidney cells using specific anti-LIMP monoclonal antibodies. Immunoelectron microscopy studies revealed the presence of LIMPs in secondary lysosomes, Golgi cisterna, and coated and uncoated vesicles located in the trans-Golgi cisterna, area. Pulse-chase experiments recorded LIMP precursors of 27 (LIMP I), 72 (LIMP II), and 86 kDa (LIMP III) and mature LIMPs of 35-50 (LIMP I), 74 (LIMP II), and 90-100 kDa (LIMP III). Time course studies on the acquisition of endoglycosidase H resistance by LIMPs indicated that all three LIMPs moved from the site of their synthesis in the endoplasmic reticulum to the medial Golgi within 30-60 min after their synthesis. All three LIMPs were fully glycosylated before leaving the Golgi system, the process during which LIMP I was retained in the trans side of the organelle. LIMP I reached the lysosomes with a halftime of 2 h and LIMPs II and III with half-times of 1 h after their synthesis by a mechanism that was independent of N-linked carbohydrates. LIMPs free of N-linked carbohydrates displayed much shorter half-lives than fully glycosylated LIMPs, suggesting an important role of the sugars in protecting LIMPs against proteolytic degradation. Double immunofluorescence microscopy experiments showed that LIMP I, LIMP II, and LIMP III are localized in the same lysosomes.
D000818:Animals; D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex / Q000032:analysis; D001692:Biological Transport; D002460:Cell Line; D006023:Glycoproteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism* / Q000648:ultrastructure; D008099:Liver / Q000378:metabolism; D008247:Lysosomes / Q000378:metabolism* / Q000648:ultrastructure; D008565:Membrane Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D008854:Microscopy, Electron; D011499:Protein Processing, Post-Translational; D051381:Rats
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D006023:Glycoproteins; D008565:Membrane Proteins