The surface activity of the same subpopulation of galactosyl receptors on isolated rat hepatocytes is modulated by colchicine, monensin, ATP depletion, and chloroquine.
In this study, we characterized and compared the ligand-independent loss of surface galactosyl (Gal) receptor activity on isolated rat hepatocytes treated with monensin, chloroquine, microtubule depolymerizing agents, or NaN3 and NaF at 37 degrees C. Freshly isolated hepatocytes exhibit predominately one subset of surface Gal receptors, termed State 1 receptors (Weigel, P. H., Clarke, B. L., and Oka, J. A. (1986) Biochem. Biophys. Res. Commun. 140, 43-50). During equilibration at 37 degrees C, these cells also express a second subset of Gal receptors at the surface, termed State 2 receptors, and routinely double their total surface Gal receptor activity. Following equilibration at 37 degrees C and then inhibitor treatment, hepatocytes bound 40-60% less 125I-asialoorosomucoid (ASOR) at 4 degrees C than did untreated cells. Treated cells maintained a basal nonmodulated level of surface receptor activity regardless of temperature, perturbant concentration, or incubation time. Loss of surface Gal receptor activity on cells treated with multiple inhibitors simultaneously or sequentially was not additive. Thus, all treatments affected the same subpopulation of surface Gal receptors. None of these inhibitors decreased surface State 1 Gal receptor activity, but all prevented the normal appearance of State 2 Gal receptors on freshly isolated cells during incubation at 37 degrees C. The endocytic capability of residual surface State 1 Gal receptors on inhibitor-treated cells varied depending on the inhibitor. Hepatocytes treated first at 24 degrees C or with colchicine at 37 degrees C internalized greater than 85% of surface-bound 125I-ASOR. In contrast, monensin- or chloroquine-treated cells internalized approximately 50% of surface-bound 125I-ASOR. Azide-treated cells internalized less than 20% of surface-bound 125I-ASOR. We conclude that only surface State 2 Gal receptor activity is sensitive to these various perturbants. State 1 Gal receptor activity is not modulated. These data are consistent with the conclusion that only State 2 Gal receptors constitutively recycle.
McAbee
D D
DD
Department of Human Biological Chemistry and Genetics, University of Texas Medical Branch, Galveston 77550.
Clarke
B L
BL
Oka
J A
JA
Weigel
P H
PH
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D001212:Asialoglycoproteins; D002738:Chloroquine / Q000494:pharmacology*; D003078:Colchicine / Q000494:pharmacology*; D004705:Endocytosis / Q000187:drug effects; D066298:In Vitro Techniques; D007700:Kinetics; D008099:Liver / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008985:Monensin / Q000494:pharmacology*; D009961:Orosomucoid / Q000031:analogs & derivatives / Q000378:metabolism; D051381:Rats; D011956:Receptors, Cell Surface / Q000187:drug effects* / Q000378:metabolism
D001212:Asialoglycoproteins; D009961:Orosomucoid; D011956:Receptors, Cell Surface; C017030:asialoorosomucoid; C039707:galactose receptor; D002738:Chloroquine; D000255:Adenosine Triphosphate; D008985:Monensin; D003078:Colchicine