The time-dependent distribution of 125I-asialo-orosomucoid-horseradish peroxidase and 131I-immunoglobulin A among three endosomal subfractions isolated from rat liver.
Three discrete endosomal fractions showing a time-dependent uptake of radioactive ligand were partially purified from rat liver. The 3,3'-diaminobenzidine (DAB)-induced density-shift protocol of Courtoy, Quintart & Baudhuin [(1984) J. Cell Biol. 98, 870-876] was used to study the distribution among these three endosomal fractions of two ligands with different intracellular destinations. Rats received both 125I-asialo-orosomucoid-horseradish peroxidase (125I-ASOR-HRP) and 131I-dIgA simultaneously by intraportal injection. The liver was fractionated at various times after injection, the three ligand-containing endosomal fractions (A, B and C) were separated and each was subjected separately to the DAB-induced density-shift procedure in which only vesicles containing 125I-ASOR-HRP are increased in density. Information on whether 131I-dIgA was co-localized or segregated from 125I-ASOR-HRP was obtained. The two ligands in the A fraction were partly segregated and partly co-localized, and this distribution appeared to be relatively unchanged with time. The two ligands in the B fraction were co-localized at all times studied. We have tentatively identified the B fraction as a compartment in which vesicle fusion has occurred. The two ligands in the C fraction were also partly co-localized and partly segregated, but the 131I-dIgA became increasingly segregated with time. This represents the first report of the purification of an endosomal subfraction specifically involved in the accumulation of multiple ligands.
Department of Biochemistry, Case Western Reserve University School of Medicine, Cleveland, OH 44106.
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Cooper
D015100:3,3'-Diaminobenzidine / Q000494:pharmacology; D000818:Animals; D037263:Asialoglycoprotein Receptor; D001212:Asialoglycoproteins / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D006735:Horseradish Peroxidase / Q000378:metabolism*; D007070:Immunoglobulin A / Q000378:metabolism*; D007457:Iodine Radioisotopes; D008024:Ligands; D008099:Liver / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D008854:Microscopy, Electron; D010544:Peroxidases / Q000378:metabolism*; D051381:Rats; D011971:Receptors, Immunologic / Q000378:metabolism; D013347:Subcellular Fractions / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure
D037263:Asialoglycoprotein Receptor; D001212:Asialoglycoproteins; D007070:Immunoglobulin A; D007457:Iodine Radioisotopes; D008024:Ligands; D011971:Receptors, Immunologic; C057124:asialo-orosomucoid-horseradish peroxidase conjugate; D015100:3,3'-Diaminobenzidine; D006735:Horseradish Peroxidase; D010544:Peroxidases