Ultrastructural localization of the mannose 6-phosphate receptor in rat liver.

Abstract

An affinity-purified rabbit antibody against rat liver mannose 6-phosphate receptor (MP-R) was prepared. The antibody was directed against a 215 kd-polypeptide and it recognized both ligand-occupied and free receptor . Anti-MP-R was used for immunofluorescence and immunoelectron microscopy of cryosections from rat liver. MP-R was demonstrated in all parenchymal liver cells, but not in endothelial lining cells. MP-R labeling was found at the entire plasma membrane, in coated pits and coated vesicles, in the compartment of uncoupling receptor and ligand, and in the Golgi complex . Lysosomes showed only scarce MP-R label .In double-labeling immunoelectron microscopy, MP-R co-localized with albumin in the Golgi cisternae and in secretory vesicles with lipoprotein particles. Cathepsin D was associated with MP-R in the Golgi cisternae . This finding indicates that MP-R/cathepsin D complexes traverse the Golgi complex on their way to the lysosomes . The possible involvement of CURL in lysosomal enzyme targeting is discussed .Studies on biosynthesis oflysosomal enzymes have established that these enzymes are synthesized in the rough endoplasmic reticulum where they are co-translationally directed from the ribosomes to the lumena ofthe endoplasmic reticulum (ER)' . The newly synthesized lysosomal enzymes are subjected to a series of modifications including glycosylation, phosphorylation, and other changes in the structure of the carbohydrate and protein moieties as they are transported from the ER to the Golgi complex and lysosomes (for recent review see references 1 and 2). Increasing evidence indicates that the enzymes are segregated from secretory proteins by membrane receptorsthat recognize mannose 6-phosphate residues, which are present uniquely on lysosomal enzymes . Biosynthesis of mannose 6-phosphate residues in the enzymes is catalyzed by N-acetylglucosamine-l-phosphotransferase and N-acetylglucosaminyl phosphodiesterase . These enzymes are associated with Golgi membrane fractions of higher or intermediate buoyant density, which probably correspond to the cis or intermediate cisternae of the Golgi complex (3, 4). These fractions are rich in a-mannosidase I (3, 4), (presumably 'Abbreviations used in this paper: CURL, compartment of uncoupling receptor and ligand; ER, endoplasmic reticulum ; GERL, Golgiassociated endoplasmic reticulum lysosomes; MP-R, mannose 6-phosphate receptors.

hVps41 and VAMP7 function in direct TGN to late endosome transport of lysosomal membrane proteins

“…2a). These albumin-positive vesicles were distinct from LAMP carriers by their irregular shape, dense content and lack of internal membranes 23 (Fig. 2a).…”

The insulin factory: a tour of the plant surroundings and a visit to the assembly line

“…proximally to the coated compartment, it is expected to take place in the Golgi stack. For lysosomal enzymes, the proper sorting of these proteins depends on the presence of mannose-6-phosphate residues which are recognized by corresponding receptors on the sorting compartment [15]; such receptors for mannose-6-phosphate have recently been demonstrated on the cisternae of the Golgi complex and this has been taken to imply that sorting of lysosomal enzymes can occur at this level [16,17].…”

Structure of the Golgi and Distribution of Reporter Molecules at 20°C Reveals the Complexity of the Exit Compartments

“…This concept was developed largely from immunogold labeling experiments using VSV-G and other viral spike proteins as well as several endogenous transmembrane proteins. These studies showed that all cisternae, including the trans-most, appear to be labeled (see for example, Geuze et al, 1984;Orci et al, 1987). However, individual thin sections rarely reveal the full extent of the cisternae in a given Golgi stack, because of convolution of the membranes and the presence of openings in the cisternae, such as fenestrae and "holes" (Rambourg et al, 1979;Rambourg and Clermont, 1990;Ladinsky et al, 1999).…”