LR11, a Mosaic LDL Receptor Family Member, Mediates the Uptake of ApoE-Rich Lipoproteins In Vitro

Abstract

Abstract-Since the molecular identification of the low density lipoprotein receptor (LDLR), an ever increasing number of related proteins have been discovered. These receptors belonging to the LDLR family are thought to play key roles in lipoprotein metabolism in a variety of tissues, including the arterial wall. We have discovered that the expression of a 250-kDa mosaic LDLR-related protein, which we termed LR11 for the presence of 11 LDLR ligand-binding repeats, is markedly induced in smooth muscle cells in the hyperplastic intima of animal models used for the study of atherosclerosis. Here, we demonstrate that the human LR11, when overexpressed in hamster cells, binds and internalizes 39-kDa receptor-associated protein (RAP), an in vitro ligand for all receptors belonging to the LDLR family. Furthermore, LR11 binds the apolipoprotein E (apoE)-rich lipoproteins, ␤-very low density lipoproteins (VLDLs), with a high affinity similar to that of other members, such as the LDLR and VLDL receptor. RAP and ␤-VLDL compete with each other; however, other serum lipoproteins are not able to inhibit their binding. LR11 shows specific binding of apoE-enriched HDL prepared from human cerebrospinal fluid as well as of ␤-VLDL, suggesting that the apoE content of lipoproteins is most likely important for mediating the high-affinity binding to the receptor. LR11-overexpressing cells are able to internalize and degrade the bound ␤-VLDL; these cells also show increased accumulation of cholesteryl esters when incubated with ␤-VLDL. Incubation for 48 hours with ␤-VLDL of LR11-overexpressing cells, but not of control cells, promotes the appearance of numerous intracellular lipid droplets. Taken together, LR11, a mosaic LDLR family member whose expression in smooth muscle cells is markedly induced in atheroma, has all the properties of a receptor for the endocytosis of lipoproteins, particularly for the incorporation of apoE-rich lipoproteins. Key Words: LDL receptors Ⅲ atherosclerosis Ⅲ smooth muscle cells Ⅲ ␤-VLDL R eceptors belonging to the LDL receptor (LDLR) family are thought to play key roles in lipoprotein metabolism in a variety of tissues, including the arterial wall. [1][2][3][4] Recent extensive histochemical studies have shown that receptors belonging to the LDLR family, as well as scavenger receptors mediating the incorporation of modified lipoproteins such as oxidized LDL, show marked induction of their expression during the formation of atherosclerotic lesions. [5][6][7][8][9] For instance, 1 LDLR family member, the so-called VLDL receptor (VLDLR/LR8), has been shown to be highly expressed in smooth muscle cells (SMCs), macrophages, and endothelial cells in rabbit atherosclerotic lesions. 7-9 Furthermore, VLDLR/LR8 has been shown not to be downregulated during ␤-VLDL-induced foam cell formation in vitro, and overexpression of the receptor in fibroblasts causes excessive lipid droplet accumulation in the transformed cells. 10 We and others have discovered and molecularly characterized a novel, unusually complex, and hi...

Interaction of the Cytosolic Domains of sorLA/LR11 with the Amyloid Precursor Protein (APP) and β-Secretase β-Site APP-Cleaving Enzyme

“…These results raised the question of whether sorLA could mediate the internalization of cell surface APP and thereby also reduce the processing of APP into sAPP and A␤. It has been shown that sorLA is capable of mediating endocytosis (Jacobsen et al, 2001) and binds apolipoprotein E-rich lipoproteins and mediate their uptake (Taira et al, 2001). When we investigated uptake of APP from the cell surface, however, we found that APP internalization is independent of sorLA.…”

Enhanced Expression of the LDL Receptor Family Member LR11 Increases Migration of Smooth Muscle Cells In Vitro

“…12,13 For analysis of phosphorylated extracellular signal-regulated kinase (ERK), cells were starved for 48 hours in 0.3% FBS-DMEM followed by the addition of platelet-derived growth factor (PDGF)-BB (R&D Systems) at 10 ng/mL for 10 minutes. For the analysis of cell-surface urokinasetype plasminogen activator receptor (uPAR), cells were treated with 0.5 U/mL of phosphatidylinositol-specific phospholipase C (SigmaAldrich) for 2 hours and the media recovered.…”

“…For the analysis of cell-surface urokinasetype plasminogen activator receptor (uPAR), cells were treated with 0.5 U/mL of phosphatidylinositol-specific phospholipase C (SigmaAldrich) for 2 hours and the media recovered. For inhibition assays, cells were starved for 24 hours, and the medium was changed to 10% FBS-DMEM containing receptor-associated protein (RAP) (10 g/mL), anti-LR11 antibody (1:2 dilution), 12 or apolipoprotein E (apoE; 50 g/mL, Cosmo Bio) for 24 hours. Equal amounts of membrane protein or phosphatidylinositol-specific phospholipase C (PI-PLC)-conditioned medium were subjected to SDS-PAGE under reducing (Western blot) and nonreducing (ligand blot) conditions.…”

“…Electrophoretic transfer of the proteins to polyvinylidene fluoride membranes (NEN Life Science) was performed for 2 hours on ice. For Western blotting, anti-LR11 antibody (1:2 dilution) 12 or polyclonal antibodies against rat uPAR (American Diagnostica), PDGF-␤ receptor (Upstate Biotechnology), or phosphorylated ERK 1/2 (New England Biolabs), were used. Development was performed with the electrochemiluminescence detection reagents.…”

SorLA Signaling by Regulated Intramembrane Proteolysis

“…Other ligands of SorLA include lipoproteins (12), PDGF-BB (23), and GDNF (24). In accordance with the multiple ligands, SorLA also seems to be involved in several functions including protein sorting (25), lipoprotein uptake (26), smooth muscle cell migration, and the development of atherosclerosis (27), but so far no signaling cascade has been identified. Recently, SorLA was identified to be involved in trafficking APP in neuronal cells.…”