The major subunit [rat hepatic lectin-1 (RHL-1)] of the asialoglycoprotein (ASGP) receptor mediates endocytosis of the iron-binding protein lactoferrin (Lf) by isolated rat hepatocytes, yet iron loading of cultured adult rat hepatocytes increases the binding and endocytosis of Lf while greatly inhibiting the uptake of desialylated ligand. In the present study, we determined whether the iron-induced Lf-binding site is RHL-1 and examined the nature of the iron-induced block in ASGP receptor endocytic function. Isolated rat hepatocytes increased their non-haem iron content from 70 to 470 p.p.b. following incubation with ferric ammonium citrate (⩽ 100 µg/ml). These conditions blocked internalization of 125I-asialo-orosomucoid (ASOR) by ≈ 90% but increased 125I-Lf endocytosis by 40%. ASOR and anti-RHL-1 sera blocked the binding and endocytosis of 125I-Lf on control cells but not on iron-loaded cells, indicating that the iron-induced Lf-binding site on hepatocytes is not RHL-1. Iron-loading of hepatocytes in the presence or absence of excess ASOR did not significantly alter the number of active ASGP receptors on the cell surface. In contrast, iron-loading decreased the number of active intracellular receptors by 40% and blocked the uptake of 125I-ASOR prebound to the cells by ≈ 80%. Under these conditions, we found an iron-dependent evolution of 88 and 140 kDa RHL-1-containing, β-mercaptoethanol-sensitive multimers that constituted up to 34 and 23%, respectively, of total immunodetectable RHL-1. We propose that iron-induced formation of cystinyl-linked RHL-1-containing multimers inhibits ASGP receptor movement between cell surface and interior and disrupts acylation of intracellular receptors.
Iron loading of isolated rat hepatocytes inhibits asialoglycoprotein receptor dynamics and induces formation of rat hepatic leptin-1 (RHL-1) oligomers
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Douglas D. McABEE, Yuan Yuan LING, Charles STICH; Iron loading of isolated rat hepatocytes inhibits asialoglycoprotein receptor dynamics and induces formation of rat hepatic leptin-1 (RHL-1) oligomers. Biochem J 1 May 1998; 331 (3): 719–726. doi: https://doi.org/10.1042/bj3310719
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