ATP-binding cassette, subfamily G (ABCG family)

This review summarizes the characteristics of the ATP-binding cassette, subfamily G (ABCG family), which has five members: ABCG1, ABCG2, ABCG4, ABCG5, and ABCG8. The members consist of a single ABC cassette in the amino terminal followed by six putative transmembrane domains, and to become functionally active, they form homo- or obligate heterodimers. Except for ABCG2, the members of the ABCG family play an important role in efflux transport of cholesterol. Mutations causing a loss of function of ABCG5 or ABCG8 are associated with sitosterolemia characterized by accumulation of phyto- and shellfish sterols. Unlike other members, ABCG2 is not involved in cholesterol efflux, but it exhibits broad substrate specificity to xenobiotic compounds. ABCG2 confers cancer cells resistance to anticancer drugs and plays a critical role in the pharmacokinetics of drugs in the clearance organs and tissue barriers. ABCG2 is also associated with a subpopulation phenotype of stem cells. Genetic polymorphisms of ABCG2 have been suggested to account for the interindividual differences in the pharmacokinetics of drugs.     

Mutations in the human ATP-binding cassette transporters ABCG5 and ABCG8 in sitosterolemia

Phytosterolemia or Sitosterolemia is a rare autosomal recessive disorder characterized by highly elevated plasma levels of plant sterols and cholesterol as a consequence of hyperabsorption and impaired biliary secretion of sterols. The disease is caused by mutations in two half size ATP-binding cassette transporters, ABCG5 and ABCG8. We have analyzed the genomic sequence of ABCG5 and ABCG8 in five well-characterized patients with Sitosterolemia. In the first patient we found a heterozygous mutation in exon 8 of the ABCG5 gene leading to a premature termination of the protein (Arg408Ter). This German patient is the first European showing a mutation of the ABCG5 gene. In a second patient we found a novel heterozygous mutation in exon 5 of ABCG8 (c.584T>A; Leu195Gln). Both patients were heterozygous for the identified mutation, but no mutation could be identified on the other chromosome. In three further analyzed patients we found mutations in exons 7, 9 and 11 of the ABCG8 gene, respectively, of which two result in a premature termination signal for translation products. One of these patients was compound heterozygous (Trp361Ter and Arg412Ter), the other was homozygous for Trp361Ter. The third patient was homozygous for an amino acid exchange (Gly574Arg). In conclusion this report describes one novel mutation affecting a highly conserved amino acid and two previously identified mutations in the ABCG8 gene. In addition, we identified for the first time a mutation in the ABCG5 gene of a European Sitosterolemia patient.     

Putative ATP-binding cassette transporter is essential for Brucella ovis pathogenesis in mice

Brucella ovis is a major cause of reproductive failure in sheep, which is associated with epididymitis and infertility in rams. Importantly, B. ovis is one of the few Brucella species that is not zoonotic. Due to the scarcity of studies on B. ovis infection, a murine model of infection was developed. The roles of B. ovis genes encoding a putative hemagglutinin and an ABC transporter were investigated in the mouse model. The kinetics of B. ovis infection were similar in BALB/c and C57BL/6 mice, and both strains of mice developed multifocal microgranulomas in the liver and spleen, but only minimal colonization and histopathological changes were observed in the genital tract. Therefore, the mouse was considered a suitable infection model for B. ovis but not for B. ovis-induced genital disease. Two mutant strains were generated in this study (the ΔabcAB and Δhmg strains). The B. ovis ΔabcAB strain was attenuated in the spleens and livers of BALB/c mice compared to the wild-type (WT) strain (P < 0.001). Conversely, the Δhmg strain infected mice at the same level as WT B. ovis, suggesting that a putative hemagglutinin is not required for B. ovis pathogenesis. Additionally, the ΔabcAB strain did not survive in peritoneal macrophages, extracellularly in the peritoneal cavity, or in RAW 264.7 macrophages. Moreover, infection with the ΔabcAB strain was not lethal for male regulatory factor 1-knockout mice, whereas infection with the B. ovis WT strain was 100% lethal within 14 days postinfection. These results confirm that the predicted ABC transporter is required for the full virulence and survival of B. ovis in vivo.     

Constitutive expression of the ATP-binding cassette transporter ABCG2 enhances the growth potential of early human hematopoietic progenitors

The ATP-binding cassette transporter, ABCG2, is a molecular determinant of the side population phenotype, which is enriched for stem and progenitor cells in various nonhematopoietic and hematopoietic tissues. ABCG2 is highly expressed in hematopoietic progenitors and silenced in differentiated hematopoietic cells, suggesting a role of ABCG2 in early hematopoiesis. To test whether ABCG2 is involved in human hematopoietic development, we retrovirally transduced umbilical cord blood-derived early hematopoietic cells and analyzed hematopoiesis in vitro and in vivo. ABCG2 increased the number of clonogenic progenitors in vitro, including the most primitive colony-forming unit-granulocyte, erythroid, macrophage, megakaryocyte, by twofold (n = 14; p < .0005). Furthermore, ABCG2 induced a threefold increase in the replating capacity of primary colonies (n = 9; p < .01). In addition, ABCG2 impaired the development of CD19+ lymphoid cells in vitro. In transplanted NOD/SCID mice, the ATP-binding cassette transporter decreased the number of human B-lymphoid cells, resulting in an inversion of the lymphoid/myeloid ratio. ABCG2 enhanced the proportion of CD34+ progenitor cells in vivo (n = 4; p < .05) and enhanced the most primitive human progenitor pool, as determined by limiting dilution competitive repopulating unit assay (p < .034). Our data characterize ABCG2 as a regulatory protein of early human hematopoietic development.     

A case of Tangier disease with a novel mutation in the C-terminal region of ATP-binding cassette transporter A1

Tangier disease (TD), a rare disorder characterized by extremely low levels of high density lipoprotein cholesterol (HDL-C), is caused by mutations in the ATP-binding cassette transporter A1 (ABCA1) gene. Here, we describe a new patient with TD. The 42-year-old proband had obvious juvenile cataracts, mild hepatosplenomegaly, and an extremely low level of HDL-C (1 mg/dl), consistent with the diagnosis of TD. The proband was homozygous for a novel CTC6914-6TT --> 2203X mutation in the carboxy terminus of the ABCA1 protein. ApoAI-mediated cholesterol efflux from patient fibroblasts was markedly decreased compared to control, despite normal levels of protein expression. This indicates that this mutant protein is normally transcribed and exists as a stable product, yet is functionally defective. These results point to a critical role for the intracellular C-terminal region of the ABCA1 gene product in regulating cholesterol efflux and HDL-cholesterol levels.     

三磷酸腺苷结合盒转运体A1对糖尿病地鼠巨噬细胞内胆固醇外流的影响

目的探讨糖尿病地鼠巨噬细胞内胆固醇外流的改变,尤其是三磷酸腺苷结合盒转运体A1(ABCA1)的功能变化以及与载脂蛋白AI(apoA-I)的相互作用。方法用金黄地鼠高脂及糖尿病模型,分离纯化腹腔巨噬细胞,用外源性apoA-I及8-br-cAMP在体外干预巨噬细胞,测定干预前后巨噬细胞ABCA1表达和细胞内胆固醇含量的变化。结果细胞内胆固醇含量在糖尿病组高于高脂组和正常对照组。apoA-I和8-br-cAMP在体外均增加糖尿病组巨噬细胞ABCA1 mRNA的表达。apoA-I使各组细胞内胆固醇外流均增加,且与孵育时间成正比,胆固醇外流量糖尿病组高于高脂组和正常对照组。结论糖尿病鼠巨噬细胞内胆固醇大量沉积,可能与内源性apoA-I量和功能的改变所引起的ABCA1介导的细胞内胆固醇外流受阻有关。     

Hepatic oval cells have the side population phenotype defined by expression of ATP-binding cassette transporter ABCG2/BCRP1

Organ-specific stem cells can be identified by the side population (SP) phenotype, which is defined by the property to effectively exclude the Hoechst 33342 dye. The ATP-binding cassette transporter ABCG2/BCRP1 mediates the SP phenotype. Because hepatic oval cells possess several characteristics of stem cells, we examined whether they have the SP phenotype using the 2-acetylaminofluorene/partial hepatectomy (PH) model. Fluorescence-activated cell sorting analysis showed that a population of non-parenchymal cells containing oval cells, prepared on day 7 after PH, carried a significant number of SP cells, whereas that of non-parenchymal cells without oval cells, prepared on day 0 after PH, did not. Northern blot analysis using total liver RNA obtained on various days after PH showed that the expression of ABCG2/BCRP1 mRNA increased after PH, reaching the highest level on day 7, and then gradually decreased. This pattern of changes in the ABCG2/BCRP1 mRNA level was well correlated to that in the number of oval cells. Furthermore, in situ hybridization revealed that oval cells were the sites of expression of ABCG2/BCRP1 mRNA. These results indicate that oval cells have the SP phenotype defined by expression of ABCG2/BCRP1, suggesting that oval cells may represent stem cells in the liver.     

Formation of cholesterol-enriched structures by aberrant intracellular accumulation of ATP-binding cassette transporter A1

ATP-binding cassette transporter A1 (ABCA1) is a key transporter associated with excess cellular lipid efflux. Here, we report that in HEK293 cells ABCA1 functions in intracellular compartments along the endocytic pathway. Inhibition of ABCA1-GFP degradation with proteasome inhibitors induced the internalization of ABCA1 and the formation of intracellular round-shaped structures, designated "A1 bodies". Importantly, cholesterol was selectively accumulated in A1 bodies, and this depended on the cholesterol efflux activity of ABCA1. Treatment with either lactacystin or acetylated LDL, which reduces proteasome activity, resulted in internalization of ABCA1 in mouse peritoneal macrophages. By performing array analysis on macrophages treated with these reagents, we identified Rab4 as a key protein involved in the internalization and aberrant accumulation of ABCA1 in HEK cells. Treatment of the cells with proteasome inhibitors inhibited the degradation of Rab4, and Rab4 over-expression induced the formation of small A1 bodies. Furthermore, A1 bodies formation was substantially inhibited by silencing of the endogenous Rab4 gene. Taken together, our findings suggest that the endocytic ABCA1 possesses cholesterol efflux activity, and thus the cellular control of post-endocytic sorting, retention or recycling of functional ABCA1 in the endocytic vesicles, which is in part regulated by Rab4, is important for cholesterol metabolism in living cells.     

ATP结合盒转运体A1基因变异与冠心病的研究进展

脂质代谢异常是冠心病的发病机制之一,ATP结合盒转运子A1( ABCA1)在胆固醇的逆向转运及高密度脂蛋白生成中起重要作用.ABCA1基因变异可引起脂质代谢紊乱,促进动脉粥样硬化及冠心病的发生发展.本文就近年来对ABCA1基因变异与冠心病关联性及其参与冠心病发病机制方面的研究做一综述.     

The mitochondrial ATP-binding cassette transporter Abcb7 is essential in mice and participates in cytosolic iron-sulfur cluster biogenesis

Proteins with iron-sulfur (Fe-S) clusters participate in multiple metabolic pathways throughout the cell. The mitochondrial ABC half-transporter Abcb7, which is mutated in X-linked sideroblastic anemia with ataxia in humans, is a functional ortholog of yeast Atm1p and is predicted to export a mitochondrially derived metabolite required for cytosolic Fe-S cluster assembly. Using an inducible Cre/loxP system to delete exons 9 and 10 of the Abcb7 gene, we examined the phenotype of mice deficient in Abcb7. We found that Abcb7 was essential in extra-embryonic tissues early in gestation and that the mutant allele exhibits an X-linked parent-of-origin lethality effect. Furthermore, using X-chromosome inactivation assays and tissue-specific deletions, Abcb7 was found to be essential for the development and function of numerous other cell types and tissues. A notable exception to this was liver, where loss of Abcb7 impaired cytosolic Fe-S cluster assembly but was not lethal. In this situation, control of iron regulatory protein 1, a key cytosolic modulator of iron metabolism, which is responsive to the availability of cytosolic Fe-S clusters, was impaired and contributed to the dysregulation of hepatocyte iron metabolism. Altogether, these studies demonstrate the essential nature of Abcb7 in mammals and further substantiate a central role for mitochondria in the biogenesis of cytosolic Fe-S proteins.     

氧化修饰减弱HDL上调人脐静脉内皮细胞ABCA1表达

血脂异常是动脉粥样硬化(atherosclerosis,AS)的始动性因素.血浆高密度脂蛋白(high density lipoprotein,HDL.)在体内能被多种因素氧化,其抗AS的作用随之减弱.     

Genetic variation of the ATP-binding cassette transporter A1 and susceptibility to coronary heart disease

ATP-binding cassette transporter A1 (ABCA1) is a member of a superfamily of membrane proteins that has attracted considerable attention as a candidate gene for coronary heart disease (CHD) based on its enzyme function as a key factor in regulating plasma HDL-C and apo A-I metabolism. It has been suggested that polymorphisms in the ABCA1 gene are risk factors for CHD, but a large number of studies have reported apparently conflicting results. To investigate this inconsistency and derive a more precise estimation of the relationship, a meta-analysis of 14,040 cases and 28,607 controls from 31 published case-control studies was performed. Five potential sources of heterogeneity including ethnicity, source of control, sample size, HWE status and genotyping method of study were also assessed. Overall, significantly decreased CHD risk was associated with 219K allele of R219K polymorphism when all studies were pooled into the meta-analysis. In the subgroup analysis by ethnicity, significantly decreased risks were found in Asians and other ethnic population for the polymorphism in all genetic models; while no significant associations were found among Caucasians. When stratified by source of controls, both population and hospital based studies get consistent positive results. However, no significant results were observed for I883M polymorphism of ABCA1 in all genetic models. In conclusion, this meta-analysis suggests that K allele of ABCA1 R219K polymorphism is a protective factor associated with decreased CHD susceptibility, but these associations vary in different ethnic populations.     

自然杀伤细胞对高与低表达ABCG2的人耐药鼻咽癌细胞杀伤作用

目的:探讨自然杀伤(NK)细胞对高与低表达三磷酸腺苷(ATP)结合转运蛋白G超家族成员2(ABCG2)人多药耐药鼻咽癌CNE2/DDP细胞(ABCG2High细胞、ABCG2Low细胞)的杀伤活性差异.方法:利用免疫磁珠技术分离ABCG2High及ABCG2LowCNE2/DDP细胞、NK细胞,LDH释放测定法检测NK细胞对K562细胞、ABCG2High<及ABCG2LowCNE2/DDP细胞的杀伤活性,流式细胞技术(FCM)检测两种靶细胞NKG2D配体表达率及杀伤后靶细胞凋亡率.结果:ABCG2High、AB-CG2Low CNE2/DDP细胞分离后ABCG2表达率分别为(91.40±2.32)%、(1.70 ±0.24)%,分选后NK细胞CD3-CD16+CD56+细胞的纯度达90%以上,LDH释放测定法检测结果显示,在效靶比为10 vs 1、20 vs 1时,NK细胞对ABCG2Low细胞、ABCG2High细胞及K562细胞的杀伤率以ABCG2High曲细胞最高,凋亡率检测结果显示ABCG2HighCNE2/DDP细胞的凋亡率为(12.90±1.51)%,ABCG2LowCNE2/DDP细胞的凋亡相率为(6.13±0.85)%,NKG2D配体表达率检测结果显示AB.CG2High CNE2/DDP细胞5种NKG2D配体(MICA、MICB、ULBP1、ULBP2、ULBP3)表达率高于ABCG2LowCNE2/DDP细胞.结论:ABCG2HighCNE2/DDP细胞比ABCG2LowCNE2/DDP细胞更容易被NK细胞杀伤,其初步的机制是ABCG2HighCNE2/DDP细胞NKG2D配体表达率高于ABCG2LowCNE2/DDP细胞,导致了ABCG2HighCNE2/DDP细胞对NK的杀伤敏感性增强.     

Signatures of recent positive selection at the ATP-binding cassette drug transporter superfamily gene loci

Members of the ATP-binding cassette (ABC) superfamily of transporters have been implicated as major players in drug response. Single nucleotide polymorphisms (SNPs) in the ABC transporter genes may account for variation in drug response between individuals. Given the abundance of SNPs within the human genome, identification of functionally important SNPs is difficult. Here, we utilized signatures of recent positive selection (RPS) to identify SNPs in ABC genes that have potential functional significance by using the long-range-haplotype test to search for signatures of RPS at 18 ABC genes involved in drug transport. From the genotype data of these 18 ABC genes in four populations extracted from the HapMap database, at least one SNP in each of these genes displayed genomic signatures of RPS in at least one population. However, only 13 SNPs in 10 ABC genes from three populations retained statistical significance after Type I error reduction. The functional significance of six of these RPS SNPs, including those that failed multiple testing correction (MTC), has been reported previously. We experimentally confirmed a functional effect for two SNPs, including one that failed to show evidence of RPS after MTC. These observations suggest that Type I error reduction may inadvertently increase Type II error. Although the remaining positively selected SNPs have yet to be functionally validated, our study illustrates the feasibility of using this strategy to identify SNPs within 'adaptive' genes that may confer functional effect, prior to testing their roles in individual/population drug response variation or in complex disease susceptibility.     

Association of ATP-binding cassette transporter A1 gene polymorphisms with plasma lipid variability and coronary heart disease risk

Objective: Our study aimed to investigate the association of ABCA1 polymorphisms with plasma lipid variability and CHD risk in the Chinese Han population. Methods: 754 CHD patients and 760 controls were included in this case-control study. Three SNPs (rs363717, rs4149339, and rs4149338) in ABCA1 3’UTR and one nonsynonymous SNP (rs2230808) in ABCA1 exon 35 were selected and genotyped. The analysis of genetic data was performed using the SNPstats program and the SPSS17.0 software. Results: Significant associations were observed between SNP rs363717 and CHD risk under different genetic models before or after Bonferroni corrections (codominant model: OR = 0.70, P = 0.003 for AG vs. AA; dominant model: OR = 0.71, P = 0.003 for GG + AG vs. AA). The nonsynonymous SNP rs2230808 was associated with higher total cholesterol levels (P = 0.047). The GCC haplotype (consisting of alleles of SNPs rs363717, rs4149339, and rs4149338) was associated with a decreased risk of CHD (OR = 0.8, P = 0.027). Three ABCA1 SNPs interacted with high triglyceride levels to increase CHD risk (P values of interactions were 0.010 for rs363717, 0.010 for rs4149339, and 0.020 for rs4149338, respectively). Conclusions: Our results suggest that ABCA1 polymorphisms influence plasma lipid variability and CHD risk. ABCA1 polymorphisms could also modify the effects of plasma lipids on CHD risk.     

A promoter variant of the ATP-binding cassette transporter A1 gene alters the HDL cholesterol level in the general Japanese population

To investigate the effects of polymorphisms in the ATP-binding cassette transporter A1 (ABCA1) gene on the high-density lipoprotein cholesterol (HDL-C) level and the incidence of myocardial infarction (MI), we performed association studies. Sequence analysis identified 14 polymorphisms in the promoter region of ABCA1. After considering linkage disequilibrium, three polymorphisms in the promoter region and 11 polymorphisms from the JSNP database were determined in 1,880 subjects recruited from the Suita Study, representing the general population in Japan. We evaluated the association between the ABCA1 genotype and HDL-C level adjusted not only for standard factors, but also for genetic factors including ApoA1 and ApoE genotypes. Of the 14 polymorphisms tested, the G(–273)C (P=0.0074), C(–297)T (P=0.0195), and IMS-JST071749 (P=0.0093) polymorphisms were significantly associated with the HDL-C level in the Suita population. We could reconfirm that the G(–273)C genotype was influential in another set of subjects (P=0.0310, n=743). However, the distribution of the ABCA1 G(–273)C genotype in subjects with MI (n=598) was not different from that in the control population (n=801). These results indicate that ABCA1 G(–273)C has a significant effect on the HDL-C level in the general Japanese population, but not on the incidence of MI.     

Ghrelin对THP-1源性泡沫细胞ATP结合盒转运子A1和G1表达的调控作用

目的： 研究单核/巨噬细胞分化成为泡沫细胞过程中ghrelin对人ATP结合盒转运子A1和G1（ABCA1/ABCG1）表达的调控作用。方法: 体外培养人源单核细胞系THP-1,由佛波酯（PMA）作用使其分化为巨噬细胞,后者可在氧化低密度脂蛋白（ox-LDL）存在条件下进一步转变为泡沫细胞。巨噬细胞加入不同浓度的Ghrelin预孵2 h后再加入ox-LDL作用不同时间,油红O染色法观察细胞内脂滴含量,运用PT-PCR法检测ABCA1和ABCG1 mRNA水平,Western blotting法检测ABCA1和ABCG1蛋白表达,采用酶法,通过荧光分光光度计检测细胞内外胆固醇含量并计算胆固醇流出率。结果: Ghrelin干预可明显减少细胞内脂滴的形成,显著增加胆固醇流出率,并能显著增加单核/巨噬细胞泡沫化过程中ABCA1和ABCG1 mRNA 水平和蛋白表达,且此作用呈浓度依赖性而不呈时间依赖性。结论: Ghrelin可能通过上调ABCA1和ABCG1转录翻译水平延缓动脉粥样硬化的发生。     

Enhanced expression of ATP-binding cassette transporter A1 in non-rafts decreases the sensitivity of vascular endothelial cells to Shiga toxin

Shiga toxin (Stx) binds to globotriaosyl ceramide (Gb3) receptors on the surface of vascular endothelial cells, which is followed by Gb3-dependent endocytosis, and initiates a cascade leading to cell damage. The Gb3 receptor is localized in lipid rafts, in which cholesterol is tightly packed primarily with sphingolipids in a liquid-ordered state. Recent studies have indicated that phosphodiesterase (PDE) type 4 inhibitors enhance the expression of ATP-binding cassette 1 (ABCA1) which promotes cholesterol efflux from non-rafts at the plasma membrane. Here we report that rolipram, a PDE4 inhibitor, reduced the sensitivity to Stx2 of human umbilical vascular endothelial cells in association with increased apolipoproteinA-I (apoA-I)-mediated cholesterol efflux, and shift of some Gb3 molecules from lipid rafts into non-rafts. Although rolipram treatment did not reduce Gb3 content at the plasma membrane and Stx binding to whole cells of HUVECs, it reduced Stx2 endocytosis. Knockdown of ABCA1 by transfection with siRNA ABCA1 in vascular endothelial cells abrogated the protective effect of rolipram on Stx2-exposed cells. Our present results suggest that the expression level of ABCA1 protein is one of critical determinants of Stx sensitivity levels in vascular endothelial cells.