The roles of cubilin and megalin, two multiligand receptors, in proximal tubule function: possible implication in the progression of renal disease

Proteins that have not been retained by the glomerulus are reabsorbed in the proximal tubule by endocytosis, a process that involves binding at the apical pole of the tubule cell, vesicular internalization and subsequent lysosomal degradation. Data presented in this review indicate that the initial recognition step involves two high molecular weight proteins, megalin and cubilin, which have multiligand properties and can therefore account for the wide variety of proteins reabsorbed. Given the potential importance of transepithelial protein traffic in the induction of interstitial fibrosis, the identification of these receptors may have implications in the progression of acute or chronic renal disease and may provide a target for therapeutic intervention.     

Megalin-dependent cubilin-mediated endocytosis is a major pathway for the apical uptake of transferrin in polarized epithelia

Cubilin is a 460-kDa protein functioning as an endocytic receptor for intrinsic factor vitamin B(12) complex in the intestine and as a receptor for apolipoprotein A1 and albumin reabsorption in the kidney proximal tubules and the yolk sac. In the present study, we report the identification of cubilin as a novel transferrin (Tf) receptor involved in catabolism of Tf. Consistent with a cubilin-mediated endocytosis of Tf in the kidney, lysosomes of human, dog, and mouse renal proximal tubules strongly accumulate Tf, whereas no Tf is detectable in the endocytic apparatus of the renal tubule epithelium of dogs with deficient surface expression of cubilin. As a consequence, these dogs excrete increased amounts of Tf in the urine. Mice with deficient synthesis of megalin, the putative coreceptor colocalizing with cubilin, also excrete high amounts of Tf and fail to internalize Tf in their proximal tubules. However, in contrast to the dogs with the defective cubilin expression, the megalin-deficient mice accumulate Tf on the luminal cubilin-expressing surface of the proximal tubule epithelium. This observation indicates that megalin deficiency causes failure in internalization of the cubilin-ligand complex. The megalin-dependent, cubilin-mediated endocytosis of Tf and the potential of the receptors thereby to facilitate iron uptake were further confirmed by analyzing the uptake of (125)I- and (59)Fe-labeled Tf in cultured yolk sac cells.     

Cubilin, a multifunctional epithelial receptor: an overview

Cubilin is a 460-kDa endocytic receptor coexpressed with megalin, a multiligand receptor of the low-density lipoprotein receptor gene family, at the apical pole of epithelial cells in the renal proximal convoluted tubule, visceral yolk sac, ileum, and placenta. The structure of cubilin is unique: it lacks a transmembrane domain and requires megalin for its internalization. The accumulation of 27 interactive CUB domains provides the potential for multiple, possibly independent interactions and functions. Cubilin is involved in the intestinal absorption of vitamin B12, the catabolism of apolipoprotein A-I by the proximal convoluted tubule and more generally in renal protein reabsorption. The role of cubilin on fetomaternal interfaces is not defined but may be related to its ability to bind and internalize high density lipoproteins.     

Contribution of cubilin and amnionless to processing and membrane targeting of cubilin-amnionless complex

Cubilin is a peripheral apical membrane receptor for multiple ligands that are taken up in several absorptive epithelia. Recently, amnionless (AMN) was identified to form a functional receptor complex with cubilin. By expression in transfected polarized MDCK cells of AMN and several cubilin fragments, including a functional "mini" version of cubilin, the processing, sorting, and membrane anchoring of the complex to the apical membrane were investigated. The results show that truncation mutants, including the N-terminal domain of cubilin, did not appear at the plasma membrane but instead were retained in the endoplasmic reticulum or partially secreted into the medium. Coexpression with AMN led to efficient transport to the apical cell surface of the cubilin constructs, which included the EGF domains, and prevented release into the medium. AMN co-precipitated with cubilin and co-localized with cubilin at the apical cell surface. Apical sorting was observed for a broad set of nonoverlapping cubilin fragments without the N-terminal region, in the absence of AMN. The preference for apical sorting disappeared when glycosylation was inhibited by tunicamycin. In conclusion, it is shown that both units contribute to the processing of the cubilin-AMN complex to the apical membrane: AMN interacts with the EGF domains of cubilin and is responsible for membrane attachment and export of the complex from the endoplasmic reticulum, whereas the extracellular cubilin molecule is responsible for apical sorting of the complex in a carbohydrate-dependent manner.     

Multiligand endocytosis and congenital defects: roles of cubilin, megalin and amnionless

Cubilin and megalin are multiligand receptors that mediate uptake of extracellular ligands. Their function has extensively been studied in the kidney where they play a key role in vitamin B12 and vitamin D homeostasis. Amnionless is a plasma membrane protein that binds to cubilin in various epithelia; the interaction cubilin-amnionless in the gut is crucial for dietary vitamin B12 uptake. Studies in patients with gene defects in these receptors, and animal models with inactivated cubilin, megalin or amnionless suggest an important role in embryonic development and normal growth. In this review we will summarize recent data on the biological function of these receptors and focus on their implication in embryonic nutrition and central nervous system malformations.     

The tandem endocytic receptors megalin and cubilin are important proteins in renal pathology

The molecular mechanisms controlling proximal tubule reabsorption of proteins have been much elucidated in recent years. Megalin and cubilin constitute two important endocytic receptor proteins involved in this process. Although structurally very different the two receptor proteins interact to mediate the reabsorption of a large number of filtered proteins, including carrier proteins important for transport and cellular uptake of several vitamins, lipids and other nutrients. Dysfunction of either protein results in tubular proteinuria and is associated with specific changes in vitamin metabolism due to the defective proximal tubular reabsorption of carrier proteins. Additional focus on the two receptors is attracted by the possible pathogenic role of excessive tubular protein uptake during conditions of increased filtration of proteins, and by recent findings implicating members of the low density lipoprotein-receptor family, which includes megalin, in the transduction of signals by association with cytoplasmic proteins.     

Genetic evidence of an accessory activity required specifically for cubilin brush-border expression and intrinsic factor-cobalamin absorption.

Cubilin is a high molecular weight multiligand receptor that mediates intestinal absorption of intrinsic factor-cobalamin and selective protein reabsorption in renal tubules. The genetic basis of selective intestinal cobalamin malabsorption with proteinuria was investigated in a canine model closely resembling human Imerslund-Gr?sbeck syndrome caused by cubilin mutations. Canine CUBN cDNA was cloned and sequenced, showing high identity with human and rat CUBN cDNAs. An intragenic CUBN marker was identified in the canine family and used to test the hypothesis of genetic linkage of the disease and CUBN loci. Linkage was rejected, indicating that the canine disorder resembling Imerslund-Gr?sbeck syndrome is caused by defect of a gene product other than cubilin. These results imply that there may be locus heterogeneity among human kindreds with selective intestinal cobalamin malabsorption and proteinuria and that normal brush-border expression of cubilin requires the activity of an accessory protein.     

The particles of the embryonic cerebrospinal fluid: How could they influence brain development?

During brain development, the embryonic cerebrospinal fluid (E-CSF) allows brain expansion and promotes neuroepithelial cell survival, proliferation or differentiation. Previous analyses of E-CSF content have revealed a high protein concentration and the presence of membranous particles. The role of these particles in the E-CSF remains poorly investigated. In this study we showed that the E-CSF contains at least two pools of particles: lipoproteins and exosome-like particles. We showed that these two populations of particles strongly interact with neuropithelial cells via an endocytic process, which display regional specificity along the developing neural tube. Finally, we explore and discuss the possibility that these interactions may influence brain development through the regulation of morphogen and growth factor signaling transduction.     

Megalin mediates the influence of sonic hedgehog on oligodendrocyte precursor cell migration and proliferation during development

Oligodendrocyte precursor cells (OPCs) of the optic nerve are generated in the preoptic area, from where they migrate to colonize it entirely. Sonic hedgehog (Shh) induces the proliferation of these cells as well as influencing their migration, acting through its canonical receptor (Ptc-1). However, the multiligand receptor megalin (or LRP-2) is also involved in Shh-induced OPC proliferation and migration, and thus, we have evaluated the relevance of this interaction. During the stages at which Shh influences OPC development, we found megalin to be selectively expressed by optic nerve astrocytes, whereas Ptc-1 and Gli1 were found in OPCs. Indeed, this pattern of expression paralleled the rostral-caudal expression of the three Shh-related molecules during the time course of plp-dm20(+) -OPC colonization. The blockage of megalin partially abolished OPC chemoattraction and fully impaired Shh-induced proliferation. Using in vitro co-cultures of dissociated optic nerve cells, we demonstrated that Shh was internalized by astrocytes via megalin, and sufficient Shh was subsequently released to produce the biological effects on OPCs observed in the nerve. Together, these data indicate that at least part of the influence of Shh on OPCs is mediated by megalin during optic nerve development, and that astrocytes expressing megalin transiently capture Shh to present it to OPCs and/or to control the gradient of this molecule during development.