125I-Insulin internalization by perfused rat liver: comparison of its subcellular distribution with that of a lysosomally targeted molecule, 125I-asialofetuin.
The subcellular distribution of 125I-insulin in the perfused rat liver was compared with the subcellular distribution of the lysosomally targeted asialoglycoprotein, 125I-asialofetuin. The use of Percoll density gradient medium provided excellent separation of lysosomes from the subcellular membrane fractions. Following perfusion with 125I-asialofetuin, a distinct peak of TCA-precipitable radioactivity could be observed in the lysosomal region of the gradient. In contrast, the gradient distribution of TCA-precipitable radioactivity following perfusion with physiological concentrations of 125I-insulin was unimodal, the observed peak corresponding to the distribution of intracellular membrane marker enzymes. Leupeptin, an inhibitor of lysosomal proteolysis, inhibited the degradation of 125I-asialofetuin but had no effect on 125I-insulin degradation. In addition, leupeptin produced a marked increase in TCA-precipitable radioactivity in the lysosome rich region of gradients prepared from livers perfused with 125I-asialofetuin. No such effect was observed following perfusion with 125I-insulin. These findings are consistent with an initial localization of the internalized insulin molecule with the membraneous system of the liver cell rather than the lysosomal system.
D000818:Animals; D001212:Asialoglycoproteins; D060748:Fetuins; D007328:Insulin / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism; D007976:Leupeptins / Q000494:pharmacology; D008099:Liver / Q000378:metabolism* / Q000648:ultrastructure; D008247:Lysosomes / Q000201:enzymology / Q000378:metabolism*; D008297:Male; D010477:Perfusion; D051381:Rats; D011919:Rats, Inbred Strains; D013347:Subcellular Fractions / Q000201:enzymology / Q000378:metabolism*; D000509:alpha-Fetoproteins / Q000378:metabolism*
D001212:Asialoglycoproteins; D060748:Fetuins; D007328:Insulin; D007976:Leupeptins; D000509:alpha-Fetoproteins; C017029:asialofetuin; C032854:leupeptin