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.
Kennedy
G
G
Department of Biochemistry, Case Western Reserve University School of Medicine, Cleveland, OH 44106.
Cooper
C
C
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