Diacytosis of asialoglycoprotein in isolated hepatocytes is dependent on the structure of ligand and cellular distribution of the receptors.
Diacytosis, degradation and retention of 125I-labeled asialoorosomucoid (ASOR), its reduced and carboxymethylated N-terminal cyanogen bromide-cleaved fragment (RC-ASCNBr-I) and asialofetuin preloaded into isolated rat or rabbit hepatocytes for various periods of time were compared. In rat hepatocytes preloaded with a saturating concentration (3.10(-8) M) of the ligands, the proportion of the preloaded ligands distributed to degradation and diacytosis was fairly constant during 4 h of preincubation. In addition, a small portion of the preloaded ligands was neither diacytosed nor degraded, but was retained intracellularly. Diacytosis of 125I-ASOR (29%) was greater than that of either 125I-RC-ASCNBr-I (23%) or 125I-asialofetuin (15%). Diacytosis of 125I-ASOR, when preloaded in the presence of 5 microM colchicine, was significantly enhanced by 79% (increasing from 29% to 52%), whereas those of 125I-RC-ASCNBr-I and 125I-asialofetuin were not significantly altered (with average increases of 14% and 19%, respectively). The fraction of the preloaded 125I-asialofetuin (69%) and 125I-RC-ASCNBr-I (68.6%) that was degraded was slightly higher than that of 125I-ASOR (64%) and all was decreased by colchicine. The fraction of all three ligands retained by the cells was increased 2- to 4-fold by colchicine. The extents of retention of 125I-asialofetuin and 125I-ASCNBr-I were greater than that of 125I-ASOR, particularly after preloaded for more than 2 h. Preloading of the cells with ligands at a non-saturating concentration (6.5.10(-10) M) did not change these patterns of ligand distribution. Conjugation of diphtheria toxin fragment A (DTA) to ASOR or RC-ASCNBr-I also did not significantly alter the pattern of ligand distribution. In rabbit hepatocytes containing more asialoglycoprotein receptors than rat cells, 125I-ASOR was diacytosed to a greater extent (50%, -colchicine; 61%, + cholchicine) but degraded to a lesser extent (33%, -colchicine; 13%, + colchicine) than was observed in rat cells. The extent of retention of 125I-ASOR in rabbit cells was also greater than that in rat cells. A similar pattern of differences between rabbit and rat hepatocytes was observed for 125I-DTA-ASOR. These results indicate that intracellular sorting of internalized asialglycoproteins between diacytosis and degradation is dependent on both the structure of the ligand and the distribution of the cellular receptors.
Chang
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Isaac Gordon Center for Digestive Diseases and Nutrition, Genesee Hospital, Rochester, New York 14607.
Chang
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D000818:Animals; D037263:Asialoglycoprotein Receptor; D001212:Asialoglycoproteins; D002462:Cell Membrane / Q000378:metabolism; D002463:Cell Membrane Permeability / Q000187:drug effects; D002478:Cells, Cultured / Q000187:drug effects; D003078:Colchicine / Q000494:pharmacology; D003488:Cyanogen Bromide / Q000494:pharmacology; D004167:Diphtheria Toxin / Q000494:pharmacology; D060748:Fetuins; D009961:Orosomucoid / Q000031:analogs & derivatives* / Q000378:metabolism; D010446:Peptide Fragments / Q000378:metabolism / Q000494:pharmacology; D011817:Rabbits; D051381:Rats; D011971:Receptors, Immunologic / Q000032:analysis*; D000509:alpha-Fetoproteins / Q000378:metabolism
D037263:Asialoglycoprotein Receptor; D001212:Asialoglycoproteins; D004167:Diphtheria Toxin; D060748:Fetuins; D009961:Orosomucoid; D010446:Peptide Fragments; D011971:Receptors, Immunologic; D000509:alpha-Fetoproteins; C017029:asialofetuin; C017030:asialoorosomucoid; C029795:diphtheria toxin fragment A; D003488:Cyanogen Bromide; D003078:Colchicine