ABSTRACT
Cerebral accumulation of amyloid-beta (Abeta) is characteristic of Alzheimer disease and of amyloid precursor protein (APP) transgenic mice. Here, we assessed the efficacy of CI-1011, an inhibitor of acyl-coenzyme A:cholesterol acyltransferase, which is suitable for clinical use, in reducing amyloid pathology in both young (6.5 months old) and aged (16 months old) human APP transgenic mice. Treatment of young animals with CI-1011 decreased amyloid plaque load in the cortex and hippocampus and reduced the levels of insoluble Abeta40 and Abeta42 and C-terminal fragments of APP in brain extracts. In aged mice, CI-1011 specifically reduced diffuse amyloid plaques with a minor effect on thioflavin S-positive dense-core plaques. Reduced diffusible amyloid was accompanied by suppression of astrogliosis and enhanced microglial activation. Collectively, these data suggest that CI-1011 treatment reduces amyloid burden in human APP mice by limiting generation and increasing clearance of diffusible Abeta.
Subject(s)
Acetates/pharmacology , Aging/drug effects , Alzheimer Disease/pathology , Amyloid/metabolism , Brain/drug effects , Sterol O-Acyltransferase/antagonists & inhibitors , Sulfonic Acids/pharmacology , Acetamides , Acetates/therapeutic use , Alzheimer Disease/cerebrospinal fluid , Alzheimer Disease/drug therapy , Alzheimer Disease/genetics , Amyloid beta-Peptides/metabolism , Amyloid beta-Protein Precursor/genetics , Animals , Apolipoproteins E/metabolism , Brain/metabolism , Cholesterol/metabolism , Disease Models, Animal , Enzyme Inhibitors/pharmacology , Enzyme Inhibitors/therapeutic use , Enzyme-Linked Immunosorbent Assay/methods , Gliosis/drug therapy , Gliosis/etiology , Humans , Image Processing, Computer-Assisted , Liver/drug effects , Liver/metabolism , Mice , Mice, Inbred C57BL , Mice, Transgenic , Mutation/genetics , Peptide Fragments/metabolism , Presenilin-1/metabolism , Pyridines/pharmacology , Sterol O-Acyltransferase/metabolism , Sulfonamides , Sulfonic Acids/therapeutic useABSTRACT
Amyloid beta-peptide (Abeta) has a central role in the pathogenesis of Alzheimer's disease (AD). Cellular cholesterol homeostasis regulates endoproteolytic generation of Abeta from the amyloid precursor protein (APP). Previous studies have identified acyl-coenzyme A: cholesterol acyltransferase (ACAT), an enzyme that regulates subcellular cholesterol distribution, as a potential therapeutic target for AD. Inhibition of ACAT activity decreases Abeta generation in cell- and animal-based models of AD through an unknown mechanism. Here we show that ACAT inhibition retains a fraction of APP molecules in the early secretory pathway, limiting the availability of APP for secretase-mediated proteolytic processing. ACAT inhibitors delayed the trafficking of immature APP molecules from the endoplasmic reticulum (ER) as shown by metabolic labeling and live-cell imaging. This resulted in partial ER retention of APP and enhanced ER-associated degradation of APP by the proteasome, without activation of the unfolded protein response pathway. The ratio of mature APP to immature APP was reduced in brains of mice treated with ACAT inhibitors, and strongly correlated with reduced brain APP-C99 and cerebrospinal fluid Abeta levels in individual animals. Our results identify a novel ACAT-dependent mechanism that regulates secretory trafficking of APP, likely contributing to decreased Abeta generation in vivo.
Subject(s)
Amyloid beta-Protein Precursor/metabolism , Sterol O-Acyltransferase/antagonists & inhibitors , Acetamides , Acetates/pharmacology , Amyloid beta-Peptides/metabolism , Animals , CHO Cells , Cricetinae , Cricetulus , Humans , Mice , Mice, Transgenic , Protein Transport/drug effects , Pyridines/pharmacology , Secretory Pathway , Sulfonamides , Sulfonic Acids/pharmacologyABSTRACT
Alzheimer disease-associated beta-amyloid peptide is generated from its precursor protein APP. By using the yeast two-hybrid assay, here we identified HtrA2/Omi, a stress-responsive chaperone-protease as a protein binding to the N-terminal cysteinerich region of APP. HtrA2 coimmunoprecipitates exclusively with immature APP from cell lysates as well as mouse brain extracts and degrades APP in vitro. A subpopulation of HtrA2 localizes to the cytosolic side of the endoplasmic reticulum (ER) membrane where it contributes to ER-associated degradation of APP together with the proteasome. Inhibition of the proteasome results in accumulation of retrotranslocated forms of APP and increased association of APP with HtrA2 and Derlin-1 in microsomal membranes. In cells lacking HtrA2, APP holoprotein is stabilized and accumulates in the early secretory pathway correlating with elevated levels of APP C-terminal fragments and increased Abeta secretion. Inhibition of ER-associated degradation (either HtrA2 or proteasome) promotes binding of APP to the COPII protein Sec23 suggesting enhanced trafficking of APP out of the ER. Based on these results we suggest a novel function for HtrA2 as a regulator of APP metabolism through ER-associated degradation.
Subject(s)
Mitochondrial Proteins/physiology , Serine Endopeptidases/physiology , Amyloid/metabolism , Animals , Brain/metabolism , CHO Cells , Cricetinae , Cricetulus , Cytosol/metabolism , Endoplasmic Reticulum/metabolism , High-Temperature Requirement A Serine Peptidase 2 , Humans , Membrane Proteins/biosynthesis , Mice , Mitochondrial Proteins/metabolism , Protein Structure, Tertiary , Serine Endopeptidases/metabolism , Subcellular Fractions/metabolism , Vesicular Transport Proteins/physiologyABSTRACT
Drosophila rely entirely on an innate immune response to combat microbial infection. Diaminopimelic acid-containing peptidoglycan, produced by Gram-negative bacteria, is recognized by two receptors, PGRP-LC and PGRP-LE, and activates a homolog of transcription factor NF-kappaB through the Imd signaling pathway. Here we show that full-length PGRP-LE acted as an intracellular receptor for monomeric peptidoglycan, whereas a version of PGRP-LE containing only the PGRP domain functioned extracellularly, like the mammalian CD14 molecule, to enhance PGRP-LC-mediated peptidoglycan recognition on the cell surface. Interaction with the imd signaling protein was not required for PGRP-LC signaling. Instead, PGRP-LC and PGRP-LE signaled through a receptor-interacting protein homotypic interaction motif-like motif. These data demonstrate that like mammals, drosophila use both extracellular and intracellular receptors, which have conserved signaling mechanisms, for innate immune recognition.