Monensin inhibits recycling of macrophage mannose-glycoprotein receptors and ligand delivery to lysosomes

Abstract

Binding studies with cells that had been permeabilized with saponin indicate that alveolar macrophages have an intracellular pool of mannose-specific binding sites which is about 4-fold greater than the cell surface pool. Monensin, a carboxylic ionophore which mediates proton movement across membranes, has no effect on binding of ligand to macrophages but blocks receptor-mediated uptake of 125I-labelled beta-glucuronidase. Inhibition of uptake was concentration- and time-dependent. Internalization of receptor-bound ligand, after warming to 37 degrees C, was unaffected by monensin. Moreover, internalization of ligand in the presence of monensin resulted in an intracellular accumulation of receptor-ligand complexes. The monensin effect was not dependent on the presence of ligand, since incubation of macrophages with monensin at 37 degrees C without ligand resulted in a substantial decrease in cell-surface binding activity. However, total binding activity, measured in the presence of saponin, was much less affected by monensin treatment. Removal of monensin followed by a brief incubation at pH 6.0 and 37 degrees C, restored both cell-surface binding and uptake activity. Fractionation experiments indicate that ligands enter a low-density (endosomal) fraction within the first few minutes of uptake, and within 20 min transfer to the lysosomal fraction has occurred. Monensin blocks the transfer from endosomal to lysosomal fraction. Lysosomal pH, as measured by the fluorescein-dextran method, was increased by monensin in the same concentration range that blocked ligand uptake. The results indicate that monensin blockade of receptor-mediated endocytosis of mannose-terminated ligands by macrophages is due to entrapment of receptor-ligand complexes and probably receptors in the pre-lysosomal compartment. The inhibition is linked with an increase in the pH of acid intracellular vesicles.

Oligolysine-based Oligosaccharide Clusters

“…Thus, although glycoclusters endocytosed by the mannose receptor end up in very acidic vesicles, glycoclusters taken up by DC-SIGN become localized in more neutral compartments. It is well known that mannose receptor-glycoconjugate complexes dissociate in early endosomes and that the released glycoconjugate is targeted to lysosomes while the receptor recycles to the cell surface (40). In contrast, our results indicate that glycoconjugates taken up by DC-SIGN might not reach the lysosomes, or at least not as quickly as those taken up by the mannose receptor.…”

Regulation of mannose receptor synthesis and turnover in mouse J774 macrophages

“…Bovine serum albumin (BSA, Fraction V, Sigma) was mannosylated [22] and iodinated using chloramine T as described [23]. Na 125 I and 14 C-methylated protein standards were purchased from Amersham Corp. Trans 35 S was obtained from ICN Biochemicals.…”

Constitutive Endocytic Recycling and Protein Kinase C-mediated Lysosomal Degradation Control KATP Channel Surface Density

“…1B). Likewise, pretreatment of the cells with monensin (35) or primaquine (36), agents that block recycling by preventing acidification of endosomes, also prevented recycling (Fig. 1B).…”