Abstract

BackgroundMembers of the low-density lipoprotein (LDL) receptor family are involved in endocytosis and in transducing signals, but also in amyloid precursor protein (APP) processing and β-amyloid secretion. ApoER2/LRP8 is a member of this family with key roles in synaptic plasticity in the adult brain. ApoER2 is cleaved after the binding of its ligand, the reelin protein, generating an intracellular domain (ApoER2-ICD) that modulates reelin gene transcription itself. We have analyzed whether ApoER2-ICD is able to regulate the expression of other LDL receptors, and we focused on LRP3, the most unknown member of this family. We analyzed LRP3 expression in middle-aged individuals (MA) and in cases with Alzheimer’s disease (AD)-related pathology, and the relation of LRP3 with APP.MethodsThe effects of full-length ApoER2 and ApoER2-ICD overexpression on protein levels, in the presence of recombinant reelin or Aβ42 peptide, were evaluated by microarray, qRT-PCRs, and western blots in SH-SY5Y cells. LRP3 expression was analyzed in human frontal cortex extracts from MA subjects (mean age 51.8±4.8 years) and AD-related pathology subjects [Braak neurofibrillary tangle stages I–II, 68.4±8.8 years; III–IV, 80.4 ± 8.8 years; V–VI, 76.5±9.7 years] by qRT-PCRs and western blot; LRP3 interaction with other proteins was assessed by immunoprecipitation. In CHO cells overexpressing LRP3, protein levels of full-length APP and fragments were evaluated by western blots. Chloroquine was employed to block the lysosomal/autophagy function.ResultsWe have identified that ApoER2 overexpression increases LRP3 expression, also after reelin stimulation of ApoER2 signaling. The same occurred following ApoER2-ICD overexpression. In extracts from subjects with AD-related pathology, the levels of LRP3 mRNA and protein were lower than those in MA subjects. Interestingly, LRP3 transfection in CHO-PS70 cells induced a decrease of full-length APP levels and APP-CTF, particularly in the membrane fraction. In cell supernatants, levels of APP fragments from the amyloidogenic (sAPPα) or non-amyloidogenic (sAPPβ) pathways, as well as Aβ peptides, were drastically reduced with respect to mock-transfected cells. The inhibitor of lysosomal/autophagy function, chloroquine, significantly increased full-length APP, APP-CTF, and sAPPα levels.ConclusionsApoER2/reelin signaling regulates LRP3 expression, whose levels are affected in AD; LRP3 is involved in the regulation of APP levels.

Highlights

  • The members of the family of low-density lipoprotein (LDL) receptors are endocytic receptors that mediate the uptake of lipoproteins and have been classically studied for their role in cholesterol transport and metabolism

  • ApoER2 overexpression increases the expression of LDL-related protein 3 (LRP3) SH-SY5Y cells were transfected with full-length ApoER2, and after 48 h, a microarray was performed

  • We focused on the analysis of LDL receptors and apolipoprotein-related genes (Table 2)

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Summary

Introduction

The members of the family of low-density lipoprotein (LDL) receptors are endocytic receptors that mediate the uptake of lipoproteins and have been classically studied for their role in cholesterol transport and metabolism. Several members of the LDL receptor family are able to modulate the amyloid precursor (APP) proteolytic processing, either by regulation of the generation of the β-amyloid peptide (Aβ) or through Aβ clearance [10,11,12,13]. ApoER2 interaction with its ligand, the reelin protein, drives to a sequential proteolytic processing, resulting in the cleavage of the receptor by α-secretase, which generates a membrane-tethered C-terminal fragment (ApoER2-CTF), followed by the cleavage by γ-secretase. Members of the low-density lipoprotein (LDL) receptor family are involved in endocytosis and in transducing signals, and in amyloid precursor protein (APP) processing and β-amyloid secretion. We have analyzed whether ApoER2-ICD is able to regulate the expression of other LDL receptors, and we focused on LRP3, the most unknown member of this family. We analyzed LRP3 expression in middle-aged individuals (MA) and in cases with Alzheimer’s disease (AD)-related pathology, and the relation of LRP3 with APP

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