Quantitative role of LAL, NPC2, and NPC1 in lysosomal cholesterol processing defined by genetic and pharmacological manipulations

Charina M Ramirez,Benny Liu,Amal Aqul,Anna M Taylor,Joyce J Repa,Stephen D Turley,John M Dietschy

doi:10.1194/jlr.m013789

Charina M Ramirez, Benny Liu + Show 5 more

Open Access

https://doi.org/10.1194/jlr.m013789

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Abstract

Lipoprotein cholesterol taken up by cells is processed in the endosomal/lysosomal (E/L) compartment by the sequential action of lysosomal acid lipase (LAL), Niemann-Pick C2 (NPC2), and Niemann-Pick C1 (NPC1). Inactivation of NPC2 in mouse caused sequestration of unesterified cholesterol (UC) and expanded the whole animal sterol pool from 2,305 to 4,337 mg/kg. However, this pool increased to 5,408 and 9,480 mg/kg, respectively, when NPC1 or LAL function was absent. The transport defect in mutants lacking NPC2 or NPC1, but not in those lacking LAL, was reversed by cyclodextrin (CD), and the ED₅₀ values for this reversal varied from ~40 mg/kg in kidney to >20,000 mg/kg in brain in both groups. This reversal occurred only with a CD that could interact with UC. Further, a CD that could interact with, but not solubilize, UC still overcame the transport defect. These studies showed that processing and export of sterol from the late E/L compartment was quantitatively different in mice lacking LAL, NPC2, or NPC1 function. In both npc2(-/-) and npc1(-/-) mice, the transport defect was reversed by a CD that interacted with UC, likely at the membrane/bulk-water interface, allowing sterol to move rapidly to the export site of the E/L compartment.

Highlights

Lipoprotein cholesterol taken up by cells is processed in the endosomal/lysosomal (E/L) compartment by the sequential action of lysosomal acid lipase (LAL), Niemann-Pick C2 (NPC2), and Niemann-Pick C1 (NPC1)
Initial experiments addressed the question of whether the lysosomal export defect was quantitatively similar in mice lacking any one of the proteins involved in sterol metabolism in the late E/L compartment
Both HP-␤-CD and SBE7-␤-CD must be exerting their effects by interacting with unesterified cholesterol (UC) at the interface between membranes in the late E/L compartment and the bulk-phase water within this compartment. These studies provide the first quantitative comparison of the defects in lysosomal cholesterol processing that follow mutations inactivating LAL, NPC2, and NPC1, and give insights into how cyclodextrin may overcome these defects and prevent disease of the target organs. All cells express these three critical proteins, and all organs of the body are continuously taking up cholesteryl ester (CE) and/or UC associated with LDL, the remnants of VLDL and chylomicrons, and, in the central nervous system (CNS), apolipoprotein E [32, 33]

Summary

Introduction

Lipoprotein cholesterol taken up by cells is processed in the endosomal/lysosomal (E/L) compartment by the sequential action of lysosomal acid lipase (LAL), Niemann-Pick C2 (NPC2), and Niemann-Pick C1 (NPC1). The transport defect in mutants lacking NPC2 or NPC1, but not in those lacking LAL, was reversed by cyclodextrin (CD), and the ED50 values for this reversal varied from ‫ف‬40 mg/kg in kidney to >20,000 mg/kg in brain in both groups This reversal occurred only with a CD that could interact with UC. A CD that could interact with, but not solubilize, UC still overcame the transport defect These studies showed that processing and export of sterol from the late E/L compartment was quantitatively different in mice lacking LAL, NPC2, or NPC1 function. In both npc2Ϫ/Ϫ and npc1Ϫ/Ϫ mice, the transport defect was reversed by a CD that interacted with UC, likely at the membrane/bulk-water interface, allowing sterol to move rapidly to the export site of the E/L compartment.—Ramirez, C.

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