ABCMdb

ABCA1 p.Cys1660Arg

Predicted by SNAP2:	A: N (53%), D: D (80%), E: D (75%), F: D (75%), G: D (75%), H: D (80%), I: D (80%), K: D (85%), L: D (75%), M: D (75%), N: D (63%), P: D (80%), Q: D (80%), R: D (80%), S: N (61%), T: D (66%), V: D (75%), W: D (85%), Y: D (80%),
Predicted by PROVEAN:	A: D, D: D, E: D, F: D, G: D, H: D, I: D, K: D, L: D, M: D, N: D, P: D, Q: D, R: D, S: D, T: D, V: D, W: D, Y: D,

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Publications
[hide] A novel missense mutation of ABCA1 in transmembran... Atherosclerosis. 2009 Sep;206(1):216-22. Epub 2009 Feb 25. Maekawa M, Kikuchi J, Kotani K, Nagao K, Odgerel T, Ueda K, Kawano M, Furukawa Y, Sakurabayashi I
A novel missense mutation of ABCA1 in transmembrane alpha-helix in a Japanese patient with Tangier disease.
Atherosclerosis. 2009 Sep;206(1):216-22. Epub 2009 Feb 25., [PMID:19344898]

Abstract [show]
Tangier disease (TD) is a hereditary disorder characterized by the severe deficiency or absence of high-density lipoprotein cholesterol (HDL-C). TD is caused by mutations in the ATP-binding cassette transporter A1 (ABCA1) gene, most of which are located in the extracellular loops and nucleotide-binding domains. Here we describe the first case of TD carrying a missense mutation in a transmembrane alpha-helix of ABCA1. A 31-year-old Japanese woman had an extremely low level of HDL-C (1mg/dl) and yellowish tonsillar swelling, leading to the diagnosis of TD. The proband was homozygous for a point mutation of T4978C in exon 37, which results in the substitution of cysteine-1660 to arginine (C1660R) in the 8th transmembrane segment of ABCA1. Her parents, grandmother, and brother were found to be heterozygous for the same mutation. Both peripheral blood leukocytes from the patient and HEK293 cells transfected with T4978C-mutated ABCA1 normally expressed ABCA1 on the plasma membrane and had normal apolipoprotein A-I-binding ability. However, apolipoprotein A-I-mediated efflux of cholesterol and phospholipids was markedly diminished in HEK293 cells transfected with T4978C-mutated ABCA1. These results suggest that this mutant is normally translated and exists as a stable product with normal localization, yet is functionally defective. Cysteine-1660 appears to be a critical residue for cholesterol transport of ABCA1.

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Sentences [show]
No. Sentence Comment
4 The proband was homozygous for a point mutation of T4978C in exon 37, which results in the substitution of cysteine-1660 to arginine (C1660R) in the 8th transmembrane segment of ABCA1. Login to comment
35 Here we report a novel mutation of ABCA1 at T4978C, which causes substitution of cysteine-1660 to arginine (C1660R), in a Japanese patient with TD. Login to comment
47 Expression vector for mutant ABCA1 carrying C1660R (pcDNA3.1-C1660R ABCA1) was generated by PCR-based site-directed mutagenesis using pcDNA3.1-wild-type ABCA1 as a template. Login to comment
72 Expression and localization of the C1660R mutant. Login to comment
73 Expression and localization of ABCA1 protein were examined by confocal microscopy in (A) HEK293 cells transfected with empty pcDNA3.1 vector (mock), pcDNA3.1-wild-type ABCA1 (WT), and pcDNA3.1-C1660R ABCA1 (C1660R) and in (B) primary PB-MNC from the patient with TD (patient), her parents (father and mother), and a healthy volunteer (normal). Login to comment
87 As shown in Supplementary Fig. 1, the patient was found to be homozygous for a T to C transition at nucleotide 4978 (codon 1660, exon 37), which results in substitution of cysteine to arginine (C1660R). Login to comment
91 Apo A-I-binding ability of the C1660R mutant. Login to comment
98 (B) Alexa 488-conjugated apo A-I binding was analyzed in HEK293 cells transfected with empty pcDNA3.1 vector (mock), pcDNA3.1-wild-type ABCA1 (WT), and pcDNA3.1-C1660R mutant (C1660R) by flow cytometry. Login to comment
104 Apo A-I-mediated cholesterol efflux of the C1660R mutant. Login to comment
105 The contents of cholesterol (A) and phospholipids (B) in the medium of HEK293 cells transiently transfected with empty pcDNA3.1 vector (mock), pcDNA3.1-wild-type ABCA1 (WT), and pcDNA3.1-C1660R mutant (C1660R) were measured after 24-h incubation in the presence (black bars) or absence (white bars) of 10 ␮g/ml apo A-I. Login to comment
114 Expression and localization of the C1660R mutant Previous studies have shown that intracellular localization of ABCA1 protein is altered by specific mutations, whereas ABCA1 is normally localized on the plasma membrane [18,19]. Login to comment
117 First, we analyzed the expression of ABCA1 in HEK293 cells transfected with mock, wild-type ABCA1, and C1660R-mutant ABCA1 expression vectors using immunofluorescence confocal microscopy. Login to comment
118 As shown in Fig. 2A, both wild-type and C1660R-mutant ABCA1 transfectants expressed ABCA1 mainly on the cell surface, whereas ABCA1 was not detected in a mock transfectant because of the absence of endogenous ABCA1 expression in HEK293 cells. Login to comment
120 A strong signal was detected only on the cell surface in the patient as well as her parents and a healthy volunteer, suggesting that the C1660R mutation does not affect the localization of ABCA1 (Fig. 2B). Login to comment
122 Apo A-I-binding ability of the C1660R mutant Next, we hypothesized that the C1660R mutant lacked apo A-I binding due to disruption of the binding sites or alterations of the conformation necessary for interaction with apo A-I. Login to comment
134 Again, there was no significant difference in apo A-I binding between wild-type and C1660R-mutant transformants (Fig. 3B), although the expression level was slightly lower in C1660R-mutant ABCA1 (Fig. 3C and D). Login to comment
135 Overall, these results suggest that the C1660R mutant retains apo A-I-binding ability. Login to comment
137 Cellular lipid efflux of the C1660R mutant Finally, we tested whether C1660R-mutant ABCA1 could efflux intracellular lipids in an apo A-I-dependent manner. Login to comment
138 As shown in Fig. 4A, apo A-I-mediated cholesterol efflux was observed in HEK293 cells expressing wild-type ABCA1 but not in C1660R-mutant transformants. Login to comment
139 Apo A-I-mediated phospholipids efflux was also severely impaired in the C1660R mutant (Fig. 4B). Login to comment
141 These results suggest that the C1660R mutant has little or no activity of cellular lipid release. Login to comment
149 The C1660R mutation found in this study is the very first missense mutation in the transmembrane domain of ABCA1 (Fig. 5). Login to comment
150 Our analyses revealed that the expression, intracellular localization, and apo A-I-binding were not impaired, but cholesterol efflux was markedly reduced in the C1660R mutant. Login to comment
156 This may be the case with C1660R. Login to comment
157 Among five family members of the proband enrolled in the study, four (parents, grandmother, and brother) are heterozygous for C1660R mutation. Login to comment
169 The C1477R mutant can be expressed on the cell surface normally, but fails to bind to apo A-I unlike C1660R [18]. Login to comment
170 This difference may reflect the localization of the mutants: C1477R is in the extracellular loop and C1660R is in the transmembrane domain. Login to comment
171 Previous literature led to the following speculation regarding the mechanisms of defective cholesterol efflux of the C1660R mutant. Login to comment
173 Alternatively, the substitution of cysteine-1660 to arginine may directly or indirectly affect substrate recognition and/or conformational changes of ABCA1 associated with the ATPase cycle. Login to comment