Richard J. Stockert

The ability of hepatic plasma membranes to bind desialylated glycoproteins as a prelude to transport and catabolism has been reported earlier. The present study describes the purification, by affinity chromatography, of an hepatic protein which retains the characteristic binding properties associated with the membranes. The isolated material, which is watersoluble and free from lipids, has been identified as a glycoprotein in which 10 % of the dry weight consists of sialic acid, galactose, mannose, and glucosamine in a molar ratio of 1:1:2:2. The integrity of the terminal sialic acid residues and the presence of calcium were shown to be absolute requirements for binding. Physical chemical studies indicated a high degree of aggregation in the final, water-soluble preparation.

Transport of macromolecules into the cell by receptor-mediated endocytosis follows a complex series of intracellular transfers, passing through distinct environments. The asialoglycoprotein receptor is a prototype of the class of receptors that constitutively enters cells via coated pits and delivers ligand to these intracellular compartments. In addition to being a model of receptor-mediated endocytosis, the presence of the receptor on hepatocytes provides a membrane-bound active site for cell-to-cell interactions, has made possible the selective targeting of chemotherapeutic agents and foreign genes, and has also been implicated as a site mediating hepatitis B virus uptake. Regulated expression of receptor subunits and their intracellular trafficking during biosynthesis and endocytosis has provided insights into the relationship of receptor structure to its overall function. As a marker of hepatocellular differentiation, its study has uncovered a unique response to intracellular guanosine 3',5'-cyclic monophosphate and translational regulation of the receptor. In this review, an overview of these diverse findings is provided in an attempt to relate the various aspects of structure and function as they impact on receptor expression.

The physiological mechanism of clearance of the mononuclear phagocyte growth factor, colony-stimulating factor 1 (CSF-1), from the circulation of normal mice was investigated by following the fate of a trace amount of i.v. injected 125I-labeled CSF-1. Macrophages selectively cleared CSF-1 by CSF-1 receptor-mediated endocytosis and degraded the growth factor intracellularly. This manner of clearance provides a feedback control mechanism whereby the rate of macrophage production is determined by the number of mature macrophages.

Dengue viruses (DEN) cause a broad spectrum of clinical manifestations including potentially life-threatening conditions such as hemorrhagic shock syndrome and less frequently acute hepatitis with liver failure and encephalopathy. In addition, dengue viruses provide a potential model to understand the initiation of hepatocyte infection by the structurally closely related hepatitis C virus (HCV), because this virus at present cannot be grown in cell culture. Although the initial steps of viral infection are a critical determinant of tissue tropism and therefore pathogenesis, little is known about the molecular basis of binding and endocytic trafficking of DEN or of any other flavivirus. Our studies revealed that binding of radiolabeled DEN to the human hepatoma cell line HuH-7 was strictly pH dependent and substantially inhibitable by the glycosaminoglycan heparin. Ligand-blot analysis, performed as a viral overlay assay, showed two heparan sulfate (HS) containing cell-surface binding proteins resolving at 33 and 37 kd. Based on the sensitivity of unprotected virus and the viral binding site on the cell surface to trypsin, viral internalization was quantified as an increase in trypsin protected virus over time. Virus trafficking to the site of degradation was inhibited by pH dissociation of the clathrin coat and dependent on IP(3)-mediated homotypic endosomal fusion. These findings confirm the hypothesis that binding and internalization of DEN by hepatocytes are mediated primarily by HS containing proteoglycans and suggest that flaviviruses traffic the major clathrin-dependent endocytic pathway during infection.

IgA present in normal human serum reacts with the hepatic receptor specific for asialoglycoproteins as demonstrated by inhibition of receptor-mediated erythroagglutination. Inhibition is reversibly abolished by the oxidation of the galactose or N-acetylgalactosamine residues of IgA with galactose oxidase. The site of receptor recognition appears to be the O-glycosidically linked oligosaccharides present on the hinge region of the IgAI subtype of IgA. The demonstration of a specific binding, in vitro, of IgA by the hepatic receptor suggests that the uptake of polymeric IgA by the liver in vivo may be mediated by this reaction.