Differential effects of leupeptin, monensin and colchicine on ligand degradation mediated by the two asialoglycoprotein receptor pathways in isolated rat hepatocytes.
We have shown that degradation of asialo-orosomucoid (ASOR) in isolated rat hepatocytes occurs by two different intracellular pathways [Clarke, Oka & Weigel (1987) J. Biol. Chem. 262, 17384-17392] mediated by two subpopulations of cell surface galactosyl (Gal) receptors, designated State 1 or State 2 receptors. In the present study, several inhibitors were tested for their effects on ligand degradation by the State 1 or State 2 pathway. Leupeptin, monensin and chloroquine completely inhibited degradation of 125I-labelled ASOR in both pathways. Dose-response studies showed, however, that the State 2 pathway was more sensitive to leupeptin or monensin than the State 1 pathway. No differences were observed with chloroquine. For example, the onset of inhibition in the State 2 and State 1 pathways occurred at about 0.05 and 0.3 microM-leupeptin respectively, a 6-fold difference. At 3.5 microM-monensin, 125I-ASOR degradation in the State 2 pathway was completely blocked, whereas degradation in the State 1 pathway was essentially unaffected. Colchicine was observed to give the largest differential sensitivity between the two pathways. The State 2 degradation pathway was about 30-fold more sensitive to colchicine than the State 1 pathway. Lumicolchicine had no affect. The onset of inhibition of the rate of 125I-ASOR degradation in the State 2 and State 1 pathways occurred at approximately 0.1 and 3.0 microM-colchicine respectively. At very high concentrations (greater than 0.1 mM), the State 1 pathway could be completely inhibited. We conclude that intracellular 125I-ASOR processing or delivery to degradative compartments in both the State 1 and State 2 Gal receptor pathways requires low pH. Ligand delivery to the degradative compartment does not require microtubules in the State 1 pathway, consistent with the very rapid onset of degradation in this pathway. The State 2 degradation pathway does require microtubules.
Clarke
B L
BL
Department of Human Biological Chemistry and Genetics, University of Texas Medical Branch, Galveston 77550.
Weigel
P H
PH
D000818:Animals; D001212:Asialoglycoproteins; D002478:Cells, Cultured; D002738:Chloroquine / Q000494:pharmacology; D003078:Colchicine / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D005690:Galactose; D007976:Leupeptins / Q000494:pharmacology*; D008099:Liver / Q000378:metabolism*; D008297:Male; D008985:Monensin / Q000494:pharmacology*; D009842:Oligopeptides / Q000494:pharmacology*; D009961:Orosomucoid / Q000378:metabolism / Q000031:analogs & derivatives; D051381:Rats; D011919:Rats, Inbred Strains; D011956:Receptors, Cell Surface / Q000378:metabolism*
D001212:Asialoglycoproteins; D007976:Leupeptins; D009842:Oligopeptides; D009961:Orosomucoid; D011956:Receptors, Cell Surface; C017030:asialoorosomucoid; C039707:galactose receptor; D002738:Chloroquine; D008985:Monensin; C032854:leupeptin; D003078:Colchicine; D005690:Galactose