Expression and functional analyses of novel mutations of ATP-binding cassette transporter-1 in Japanese patients with high-density lipoprotein deficiency.
ATP-binding cassette transporter-1 (ABCA1) gene is mutated in patients with familial high-density lipoprotein deficiency (FHD). In order to know the molecular basis for FHD, we characterized three different ABCA1 mutations associated with FHD (G1158A/A255T, C5946T/R1851X, and A5226G/N1611D) with respect to their expression in the passaged fibroblasts from the patients and in the cells transfected with the mutated cDNAs. Fibroblasts from the all patients showed markedly decreased cholesterol efflux to apolipoprotein (apo)-Al. In the fibroblasts homozygous for G1158A/A255T, the immunoreactive mass of ABCA1 could not be detected, even when stimulated by 9-cis-retinoic acid and 22-R-hydroxycholesterol. In the fibroblasts homozygous for C5946T/R1851X, ABCA1 mRNA was comparable. Because the mutant ABCA1 protein (R1851X) was predicted to lack the epitope for the antibody used, we transfected FLAG-tagged truncated mutant (R1851X/ABCA1-FLAG) cDNA into Cos-7 cells, showing that the mutant protein expression was markedly reduced. The expression of N1611D ABCA1 protein was comparable in both fibroblasts and overexpressing cells, although cholesterol efflux from the cells was markedly reduced. These data indicated that, in the three patients investigated, the abnormalities and dysfunction of ABCA1 occurred at the different levels, providing important information about the expression, regulation, and function of ABCA1.
Nishida
Yoshiharu
Y
Department of Internal Medicine and Molecular Science, Osaka University, Osaka, Japan.
Hirano
Kenichi
K
Tsukamoto
Kosuke
K
Nagano
Makoto
M
Ikegami
Chiaki
C
Roomp
Kirsten
K
Ishihara
Mitsuaki
M
Sakane
Naoki
N
Zhang
Zhongyan
Z
Tsujii Ki
Ken-ichi
K
Matsuyama
Akifumi
A
Ohama
Tohru
T
Matsuura
Fumihiko
F
Ishigami
Masato
M
Sakai
Naohiko
N
Hiraoka
Hisatoyo
H
Hattori
Hiroaki
H
Wellington
Cheryl
C
Yoshida
Yoshihide
Y
Misugi
Susumu
S
Hayden
Michael R
MR
Egashira
Toru
T
Yamashita
Shizuya
S
Matsuzawa
Yuji
Y
D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters / Q000096:biosynthesis / Q000235:genetics*; D000368:Aged; D000077556:Alitretinoin; D000818:Animals; D016632:Apolipoprotein A-I / Q000378:metabolism; D019556:COS Cells; D002478:Cells, Cultured; D002784:Cholesterol / Q000378:metabolism; D004252:DNA Mutational Analysis; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D005260:Female; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D015870:Gene Expression; D017930:Genes, Reporter; D006720:Homozygote; D006801:Humans; D006888:Hydroxycholesterols / Q000494:pharmacology; D007564:Japan / Q000453:epidemiology; D008075:Lipoproteins, HDL / Q000172:deficiency*; D008297:Male; D008875:Middle Aged; D009154:Mutation / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism; D013631:Tangier Disease / Q000453:epidemiology / Q000235:genetics*; D014162:Transfection; D014212:Tretinoin / Q000494:pharmacology
C577836:ABCA1 protein, human; D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters; D016632:Apolipoprotein A-I; D018076:DNA, Complementary; D006888:Hydroxycholesterols; D008075:Lipoproteins, HDL; D012333:RNA, Messenger; C003585:22-hydroxycholesterol; D000077556:Alitretinoin; D014212:Tretinoin; D002784:Cholesterol