Acyl-coenzyme A:cholesterol acyltransferases Chang, Ta-Yuan; Li, Bo-Liang; Chang, Catherine C. Y ...
American journal of physiology: endocrinology and metabolism,
07/2009, Volume:
297, Issue:
1
Journal Article
Peer reviewed
Open access
1 Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire; 2 State Key Laboratory of Molecular Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for ...Biological Sciences, Chinese Academy of Sciences, Shanghai, China
Submitted 12 November 2008
; accepted in final form 12 January 2009
ABSTRACT
The enzymes acyl-coenzyme A (CoA):cholesterol acyltransferases (ACATs) are membrane-bound proteins that utilize long-chain fatty acyl-CoA and cholesterol as substrates to form cholesteryl esters. In mammals, two isoenzymes, ACAT1 and ACAT2, encoded by two different genes, exist. ACATs play important roles in cellular cholesterol homeostasis in various tissues. This chapter summarizes the current knowledge on ACAT-related research in two areas: 1 ) ACAT genes and proteins and 2 ) ACAT enzymes as drug targets for atherosclerosis and for Alzheimer's disease.
acyl-coenzyme A:cholesterol acyltransferase inhibitors; atherosclerosis; Alzheimer's disease
Address for reprint requests and other correspondence: T. Y. Chang, Dept. of Biochemistry, Dartmouth Medical School, 1 Rope Ferry Rd., Hanover, NH 03755-1404 (e-mail: Ta.Yuan.Chang{at}Dartmouth.edu )
24(S)-Hydroxycholesterol (24S-OHC), which is enzymatically produced in the brain, has been known to play an important role in maintaining cholesterol homeostasis in the brain and has been proposed as ...a possible biomarker of neurodegenerative disease. Recent studies have revealed diverse functions of 24S-OHC and gained increased attention. For example, 24S-OHC at sublethal concentrations has been found to induce an adaptive response via activation of the liver X receptor signaling pathway, thereby protecting neuronal cells against subsequent oxidative stress. It has also been found that physiological concentrations of 24S-OHC suppress amyloid-β production via downregulation of amyloid precursor protein trafficking in neuronal cells. On the other hand, high concentrations of 24S-OHC have been found to induce a type of nonapoptotic programmed cell death in neuronal cells expressing little caspase-8. Because neuronal cell death induced by 24S-OHC has been found to proceed by a unique mechanism, which is different from but in some ways similar to necroptosis—necroptosis being a type of programmed necrosis induced by tumor necrosis factor α—neuronal cell death induced by 24S-OHC has been called “necroptosis-like” cell death. 24S-OHC-induced cell death is dependent on the formation of 24S-OHC esters but not on oxidative stress. This review article discusses newly reported aspects of 24S-OHC in neuronal cell death and sheds light on the possible importance of controlling 24S-OHC levels in the brain for preventing neurodegenerative disease.
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•Cholesterol in converted to 24S-OHC by CYP46A1 in neurons within the brain.•24S-OHC plays a role in pathophysiology of neurodegenerative diseases.•24S-OHC induces apoptosis or necroptosis-like cell death depending on caspase-8.•24S-OHC-induced cell death is dependent on formation of 24S-OHC esters.•24S-OHC induces ROS-independent cell death in undifferentiated neuronal cells
PARK7/DJ-1 is a Parkinson disease- and cancer-associated protein that functions as a multifunctional protein involved in gene transcription regulation and anti-oxidative defense. Although PARK7 lacks ...the secretory signal sequence, it is secreted and plays important physiological and pathophysiological roles. Whereas secretory proteins that lack the endoplasmic reticulum-targeting signal sequence are secreted from cells by way of what is called the unconventional secretion mechanism, the specific processes responsible for causing PARK7 to be secreted across the plasma membrane have remained unclear. In the present study, we found that PARK7 secretion was increased by treatment with 6-OHDA via the unconventional secretory pathway in human neuroblastoma SH-SY5Y cells and MEF cells. We also found that 6-OHDA-induced PARK7 secretion was suppressed in Atg5-, Atg9-, or Atg16l1-deficient MEF cells or ATG16L1 knockdown SH-SY5Y cells, indicating that the autophagy-based unconventional secretory pathway is involved in PARK7 secretion. We moreover observed that 6-OHDA-derived electrophilic quinone induced oxidative stress as indicated by a decrease in glutathione levels, and that this was suppressed by pretreatment with antioxidant NAC. We further found that NAC treatment suppressed autophagy and PARK7 secretion. We also observed that 6-OHDA-induced autophagy was associated with activation of AMPK and ULK1 via a pathway which was independent of MTOR. Collectively these results suggest that electrophilic 6-OHDA quinone enhances oxidative stress, and that this is followed by AMPK-ULK1 pathway activation and induction of secretory autophagy to produce unconventional secretion of PARK7.
Abbreviations: 6-OHDA: 6-hydroxydopamine; AMPK: AMP-activated protein kinase; ATG: autophagy related; CAV1: caveolin 1; ER: endoplasmic reticulum; FN1: fibronectin 1; GSH: glutathione; IDE: insulin degrading enzyme; IL: interleukin; LDH: lactate dehydrogenase; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MEF: mouse embryonic fibroblast; MTOR: mechanistic target of rapamycin kinase; NAC: N-acetyl-L-cysteine; PARK7/DJ-1: Parkinsonism associated deglycase; PD: Parkinson disease; RPS6KB1/p70S6K: ribosomal protein S6 kinase B1; RPN1: ribophorin I; ROS: reactive oxygen species; ULK1: unc-51 like autophagy activating kinase 1; WT: wild-type
A vast body of evidence implicates increased oxidative stress and extracellular glutamate accumulation in the pathomechanism of sporadic amyotrophic lateral sclerosis (ALS). Cystine/glutamate ...antiporter (xCT) carries extracellular cystine uptake and intracellular glutamate release (cystine/glutamate exchange) in the presence of oxidative stress. The aim of the present study was to determine the involvement of xCT in ALS. Immunohistochemical observations in the spinal cord sections demonstrated that xCT was mainly expressed in astrocytes, with staining more intense in 12 sporadic ALS patients as compared to 12 age‐matched control individuals. Western blot and densitometric analyses of the spinal cord samples revealed that the relative value of xCT/β‐actin optical density ratio was significantly higher in the ALS group as compared to the control group. Next, we conducted cell culture experiments using a human astrocytoma‐derived cell line (1321N1) and a mouse motor neuron/neuroblastoma hybrid cell line (NSC34). In 1321N1 cells, the normalized xCT expression levels in cell lysates were significantly increased by H2O2 treatment. Glutamate concentrations in 1321 N1 cell culture‐conditioned media were significantly elevated by H2O2 treatment, and the H2O2‐driven elevations were completely canceled by the xCT inhibitor erastin pretreatment. In motor neuron‐differentiated NSC34 cells (NSC34d cells), both the normalized xCT expression levels in the cell lysates and glutamate concentrations in the cell‐conditioned media were constant with or without H2O2 treatment. The present results provide in vivo and in vitro evidence that astrocytes upregulate xCT expression to release glutamate in response to increased oxidative stress associated with ALS, contributing to extracellular glutamate accumulation.
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•24S-OHC induces apoptosis and necroptosis-like cell death in SH-SY5Y cells.•ROS generation is not involved in 24S-OHC-induced neuronal cell death.•24S-OHC-induced neuronal cell death ...is inhibited by α-Toc but not by α-Toc3.•Radical-madiated neuronal cell death is inhibited by either α-Toc or α-Toc3.•α-Toc inhibits 24S-OHC-induced cell death not by radical scavenging activity.
24(S)-Hydroxycholesterol (24S-OHC) has diverse physiological and pathological functions. In particular, cytotoxic effects of 24S-OHC in neuronal cells are important in development of neurodegenerative diseases. 24S-OHC induces necroptosis-like cell death in SH-SY5Y cells expressing little caspase-8. In the present study, 24S-OHC was found to induce apoptosis as determined by caspase-3 activation in all-trans-retinoic acid (atRA)-treated SH-SY5Y cells in which expression of caspase-8 was induced. 24S-OHC-induced cell death was inhibited by α-tocopherol (α-Toc) but not by α-tocotrienol (α-Toc3) in SH-SY5Y cells regardless of whether cells were treated with atRA. In contrast, cumene hydroperoxide (CumOOH)-induced cell death was significantly inhibited by α-Toc and α-Toc3. In atRA-treated SH-SY5Y cells, generation of reactive oxygen species (ROS) was induced by stimulation with CumOOH but was not induced by stimulation with 24S-OHC. These results suggest that inhibition of 24S-OHC-induced cell death by α-Toc cannot be explained by its radical scavenging antioxidant activity. Esterification of 24S-OHC followed by lipid droplet (LD) formation due to acyl-CoA:cholesterol acyltransferase 1 (ACAT1) are key events in 24S-OHC-induced cell death in atRA-treated SH-SY5Y cells as demonstrated by inhibition of cell death by ACAT1 inhibitor. LD number was not changed by treatment with either α-Toc or α-Toc3. The different physical properties of α-Toc and α-Toc3 may account for their different inhibitory effects on 24S-OHC-induced cell death.
Abstract
Perturbation of proteostasis triggers the adaptive responses that contribute to the homeostatic pro-survival response, whereas disruption of proteostasis can ultimately lead to cell death. ...Brain-specific oxysterol—i.e., 24(
S
)-hydroxycholesterol (24S-OHC)—has been shown to cause cytotoxicity when esterified by acyl-CoA:cholesterol acyltransferase 1 (ACAT1) in the endoplasmic reticulum (ER). Here, we show that the accumulation of 24S-OHC esters caused phosphorylation of eukaryotic translation initiator factor 2α (eIF2α), dissociation of polysomes, and formation of stress granules (SG), resulting in robust downregulation of global protein
de novo
synthesis in human neuroblastoma SH-SY5Y cells. We also found that integrated stress response (ISR) activation through PERK and GCN2 activation induced by 24S-OHC treatment caused eIF2α phosphorylation. 24S-OHC-inducible SG formation and cell death were suppressed by inhibition of ISR. These results show that ACAT1-mediated 24S-OHC esterification induced ISR and formation of SG, which play crucial roles in 24S-OHC-inducible protein synthesis inhibition and unconventional cell death.
The 24(S)-hydroxycholesterol (24S-OHC), which plays an important role in maintaining brain cholesterol homeostasis, has been shown to possess neurotoxicity. We have previously reported that 24S-OHC ...esterification by ACAT1 and the resulting lipid droplet (LD) formation are responsible for 24S-OHC-induced cell death. In the present study, we investigate the functional roles of 24S-OHC esters and LD formation in 24S-OHC-induced cell death, and we identify four long-chain unsaturated fatty acids (oleic acid, linoleic acid, arachidonic acid, and DHA) with which 24S-OHC is esterified in human neuroblastoma SH-SY5Y cells treated with 24S-OHC. Here, we find that cotreatment of cells with 24S-OHC and each of these four unsaturated fatty acids increases prevalence of the corresponding 24S-OHC ester and exacerbates induction of cell death as compared with cell death induced by treatment with 24S-OHC alone. Using electron microscopy, we find in the present study that 24S-OHC induces formation of LD-like structures coupled with enlarged endoplasmic reticulum (ER) lumina, and that these effects are suppressed by treatment with ACAT inhibitor. Collectively, these results illustrate that ACAT1-catalyzed esterification of 24S-OHC with long-chain unsaturated fatty acid followed by formation of atypical LD-like structures at the ER membrane is a critical requirement for 24S-OHC-induced cell death.
The present study investigated the therapeutic effects of the curcumin derivative 3-(1E)-2-(1H-indol-6-yl)ethenyl-5-(1E)-2-2-methoxy-4-(2-pyridylmethoxy)phenylethenyl-1H-pyrazole (GT863) in ...amyotrophic lateral sclerosis (ALS). The inhibitory effect of GT863 on superoxide dismutase 1 (SOD1) aggregation was evaluated in cell-free assays. GT863 interfered with the conformational changes of the SOD1 protein and later, oligomeric aggregation. Furthermore, its antioxidant, anti-inflammatory, and neuroprotective effects were evaluated in cell-free and cultured cell assays. GT863 inhibited H
2
O
2
− and glutamate-induced cytotoxicity and activated an antioxidant responsive element pathway. Additionally, in vivo effects of GT863 in the ALS mice model were evaluated by its oral administration to H46R mutant SOD1 transgenic mice. Rotarod test showed that GT863 administration significantly slowed the progression of motor dysfunction in the mice. In addition, GT863 substantially reduced highly-aggregated SOD1, further preserving large neurons in the spinal cord of GT863-treated mice. Collectively, these results indicated that GT863 could be a viable therapeutic agent with multiple vital actions for the treatment of ALS.
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We synthesized several candidates of 24(S)-hydroxycholesterol (24S-OHC) esters, which are involved in neuronal cell death, through catalysis with acyl-CoA:cholesterol ...acyltransferase-1 (ACAT-1). We studied the regioselectivity of the acylation of the secondary alcohol at the 3- or 24-position of 24S-OHC. The appropriate saturated and unsaturated long-chain fatty acids were esterified with the protected 24S-OHC and then de-protected to afford the desired esters at a satisfactory yield. We then confirmed by HPLC monitoring that the retention times of four esters of 24S-OHC, namely 3-oleate, 3-linoleate, 3-arachidonoate and 3-docosahexaenoate, were consistent with those of 24S-OHC esters observed in 24S-OHC-treated SH-SY5Y cells.
The side-chain hydroxylation of cholesterol by specific enzymes produces 24(S)-hydroxycholesterol, 25-hydroxycholesterol, 27-hydroxycholesterol, and other products. These enzymatically formed ...side-chain oxysterols act as intermediates in the biosynthesis of bile acids and serve as signaling molecules that regulate cholesterol homeostasis. Besides these intracellular functions, an imbalance in oxysterol homeostasis is implicated in pathophysiology. Furthermore, growing evidence reveals that oxysterols affect cell proliferation and cause cell death. This chapter provides an overview of the pathophysiological role of side-chain oxysterols in developing human diseases. We also summarize our understanding of the molecular mechanisms underlying the induction of various forms of cell death by side-chain oxysterols.
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