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First published online 1 March 2005
doi: 10.1242/jcs.01725

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Intracellular fate of LDL receptor family members depends on the cooperation between their ligand-binding and EGF domains

Department of Biochemical Physiology and Institute of Biomembranes, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands

^* Author for correspondence (e-mail: k.w.rodenburg{at}bio.uu.nl)

Accepted 13 January 2005

The insect low-density lipoprotein (LDL) receptor (LDLR) homologue LpR mediates endocytosis of an insect lipoprotein (lipophorin) that is structurally related to LDL. Despite these similarities, lipophorin and LDL follow distinct intracellular routes upon endocytosis by their receptors. Whereas LDL is degraded in lysosomes, lipophorin is recycled in a transferrin-like manner. We constructed several hybrid receptors composed of Locusta migratoria LpR and human LDLR regions to identify the domains implicated in LpR-mediated ligand recycling. Additionally, the triadic His562 residue of LDLR, which is putatively involved in ligand uncoupling, was mutated to Asn, corresponding to Asn643 in LpR, to analyse the role of the His triad in receptor functioning. The familial hypercholesterolaemia (FH) class 5 mutants LDLR_H562Y and LDLR_H190Y were also analysed in vitro. Fluorescence microscopic investigation and quantification suggest that LpR-mediated ligand recycling involves cooperation between the ligand-binding domain and epidermal growth factor (EGF) domain of LpR, whereas its cytosolic tail does not harbour motifs that affect this process. LDLR residue His562 appears to be essential for LDLR recycling after ligand endocytosis but not for constitutive receptor recycling. Like LDLR_H562N, LDLR_H562Y did not recycle bound ligand; moreover, the intracellular distribution of both mutant receptors after ligand incubation coincides with that of a lysosomal marker. The LDLR mutant characterization in vitro suggests that LDLR FH class 5 mutations might be divided into two subclasses.

Key words: Lipophorin, Lipoprotein, Transferrin, Receptor, Endocytosis, Recycling

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