Impaired Retromer Function in Niemann-Pick Type C Disease Is Dependent on Intracellular Cholesterol Accumulation

Abstract

Niemann-Pick type C disease (NPC) is a rare inherited neurodegenerative disorder characterized by an accumulation of intracellular cholesterol within late endosomes and lysosomes due to NPC1 or NPC2 dysfunction. In this work, we tested the hypothesis that retromer impairment may be involved in the pathogenesis of NPC and may contribute to increased amyloidogenic processing of APP and enhanced BACE1-mediated proteolysis observed in NPC disease. Using NPC1-null cells, primary mouse NPC1-deficient neurons and NPC1-deficient mice (BALB/cNctr-Npc1m1N), we show that retromer function is impaired in NPC. This is manifested by altered transport of the retromer core components Vps26, Vps35 and/or retromer receptor sorLA and by retromer accumulation in neuronal processes, such as within axonal swellings. Changes in retromer distribution in NPC1 mouse brains were observed already at the presymptomatic stage (at 4-weeks of age), indicating that the retromer defect occurs early in the course of NPC disease and may contribute to downstream pathological processes. Furthermore, we show that cholesterol depletion in NPC1-null cells and in NPC1 mouse brains reverts retromer dysfunction, suggesting that retromer impairment in NPC is mechanistically dependent on cholesterol accumulation. Thus, we characterized retromer dysfunction in NPC and propose that the rescue of retromer impairment may represent a novel therapeutic approach against NPC.

Transfection of Sponge Cells and Intracellular Localization of Cancer-Related MYC, RRAS2, and DRG1 Proteins

“…Immunocytochemistry on HeLa and MJ90 cells was performed as previously described [ 114 ]. In short, the cells grown on coverslips were washed three times in the PBS and fixed with 4% sucrose/paraformaldehyde for 15 min.…”

Structure and function of cancer-related developmentally regulated GTP-binding protein 1 (DRG1) is conserved between sponges and humans

“…Control cells were co-transfected with empty-GFP (pEGFP-N1) and empty-CHERRY (pmCherry - C1) vector, respectively. Immunocytochemistry on MCF-7 cells was performed as previously described 70 . In short, the cells grown on coverslips were washed three times in PBS, fixed with 4% sucrose/paraformaldehyde for 15 min and permeabilized with 0.2% saponin in PBS.…”

CD63 regulates cholesterol storage within endosomes and its distribution via exosomes

“…We found that CD63 KO increases the recruitment of VPS35 on endosomes. Cholesterol availability at the limiting membrane of endosomes in absence of CD63 might control actin bundling and retromer recruitment 56,57 . This process could also rely on the potential interaction between CD63 and PI4-kinase 58 and the recent implication of this family of enzyme and PI4P in endosomal tubule formation driven by actin and retromer 59,60 .…”