三磷酸腺苷结合盒转运体和细胞胆固醇外排

胆固醇是细胞质膜的重要组成成分。然而,过多的胆固醇累积可导致细胞中毒。异常的胆固醇胞内迁移与蓄积是造成许多心血管疾病如动脉粥样硬化的分子基础。细胞内胆固醇稳态由胆固醇的吸收、合成及外排等一系列过程调控。在哺乳动物细胞中,调节胆固醇合成、吸收和外排是维持体内胆固醇平衡的必要生理过程。本综述着重概述了三磷酸腺苷结合盒转运体(ABC)家族,如ABCA1、ABCG1、ABCG5和ABCG8的细胞功能及生理作用,以及这些转运体在调控胆固醇胞外转运中的分子机制。     

Lipid efflux by the ATP-binding cassette transporters ABCA1 and ABCG1

Plasma levels of high-density lipoproteins (HDL) and apolipoprotein A-I (apoA-I) are inversely correlated with the risk of cardiovascular disease. One major atheroprotective mechanism of HDL and apoA-I is their role in reverse cholesterol transport, i.e., the transport of excess cholesterol from foam cells to the liver for secretion. The ATP-binding cassette transporters ABCA1 and ABCG1 play a pivotal role in this process by effluxing lipids from foam cells to apoA-I and HDL, respectively. In the liver, ABCA1 activity is one rate-limiting step in the formation of HDL. In macrophages, ABCA1 and ABCG1 prevent the excessive accumulation of lipids and thereby protect the arteries from developing atherosclerotic lesions. However, the mechanisms by which ABCA1 and ABCG1 mediate lipid removal are still unclear. Particularly, three questions remain controversial and are discussed in this review: (1) Do apoA-I and HDL directly interact with ABCA1 and ABCG1, respectively? (2) Does cholesterol efflux involve retroendocytosis of apoA-I or HDL? (3) Which lipids are directly transported by ABCA1 and ABCG1?     

ATP-binding cassette transporters in Escherichia coli

ATP-binding cassette (ABC) transporters are integral membrane proteins that actively transport molecules across cell membranes. In Escherichia coli they consist primarily of import systems that involve in addition to the ABC transporter itself a substrate binding protein and outer membrane receptors or porins, and a number of transporters with varied functions. Recent crystal structures of a number of ATPase domains, substrate binding proteins, and full-length transporters have given new insight in the molecular basis of transport. Bioinformatics approaches allow an approximate identification of all ABC transporters in E. coli and their relation to other known transporters. Computational approaches involving modeling and simulation are beginning to yield insight into the dynamics of the transporters. We summarize the function of the known ABC transporters in E. coli and mechanistic insights from structural and computational studies.     

Scavenger receptor-BI inhibits ATP-binding cassette transporter 1- mediated cholesterol efflux in macrophages

Scavenger receptor BI (SR-BI) facilitates the efflux of cellular cholesterol to plasma high density lipoprotein (HDL). Recently, the ATP-binding cassette transporter 1 (ABC1) was identified as a key mediator of cholesterol efflux to apolipoproteins and HDL. The goal of the present study was to determine a possible interaction between the SR-BI and ABC1 cholesterol efflux pathways in macrophages. Free cholesterol efflux to HDL was increased ( approximately 2.2-fold) in SR-BI transfected RAW macrophages in association with increased SR-BI protein levels. Treatment of macrophages with 8-bromo-cAMP (cAMP) resulted in a 4.1-fold increase in ABC1 mRNA level and also increased cholesterol efflux to HDL (2.2-fold) and apoA-I (5.5-fold). However, in SR-BI transfected RAW cells, cAMP treatment produced a much smaller increment in cholesterol efflux to HDL (1.1-fold) or apoA-I (3.3-fold) compared with control cells. In macrophages loaded with cholesterol by acetyl-LDL treatment, SR-BI overexpression did not increase cholesterol efflux to HDL but did inhibit cAMP-mediated cholesterol efflux to apoA-I or HDL. SR-BI neutralizing antibody led to a dose- and time-dependent increase of cAMP-mediated cholesterol efflux in both SR-BI transfected and control cells, indicating that SR-BI inhibits ABC1-mediated cholesterol efflux even at low SR-BI expression level. Transfection of a murine ABC1 cDNA into 293 cells led to a 2.3-fold increase of cholesterol efflux to apoA-I, whereas co-transfection of SR-BI with ABC1 blocked this increase in cholesterol efflux. SR-BI and ABC1 appear to have distinct and competing roles in mediating cholesterol flux between HDL and macrophages. In nonpolarized cells, SR-BI promotes the reuptake of cholesterol actively effluxed by ABC1, creating a futile cycle.     

The role of different regions of ATP-binding cassette transporter A1 in cholesterol efflux

ABCA1 is a key element of cholesterol efflux, but the mechanism of ABCA1-dependent cholesterol efflux is still unclear. Monoclonal antibodies against ABCA1 were used to map functional domains of ABCA1. Two antibodies were directed against a fragment of the first extracellular loop of ABCA1, and the third antibody was directed against a fragment of the fourth extracellular loop. One antibody against the first loop inhibited cholesterol efflux from human macrophages without inhibiting apolipoprotein A-I (apoA-I) binding and internalization. Another antibody against the first loop inhibited apoA-I binding and internalization without inhibiting cholesterol efflux. The antibody against the fourth loop inhibited apoA-I binding to ABCA1 but enhanced cholesterol efflux from macrophages and reduced intracellular cholesterol content. This antibody also increased cholesterol efflux from HeLa cells transfected with ABCA1 but not from cells with DeltaPEST-ABCA1. The mechanism of the stimulating effect of this antibody on cholesterol efflux was found to be stabilization of ABCA1 leading to the increase in abundance of cell surface ABCA1. We conclude that a site on the first extracellular loop is required for cholesterol efflux, whereas a site on the fourth extracellular loop may be responsible for ABCA1 stability.     

ATP-binding cassette transporters in Escherichia coli

ATP-binding cassette (ABC) transporters are integral membrane proteins that actively transport molecules across cell membranes. In Escherichia coli they consist primarily of import systems that involve in addition to the ABC transporter itself a substrate binding protein and outer membrane receptors or porins, and a number of transporters with varied functions. Recent crystal structures of a number of ATPase domains, substrate binding proteins, and full-length transporters have given new insight in the molecular basis of transport. Bioinformatics approaches allow an approximate identification of all ABC transporters in E. coli and their relation to other known transporters. Computational approaches involving modeling and simulation are beginning to yield insight into the dynamics of the transporters. We summarize the function of the known ABC transporters in E. coli and mechanistic insights from structural and computational studies.     

Preferential ATP-binding cassette transporter A1-mediated cholesterol efflux from late endosomes/lysosomes

Recently, ATP-binding cassette transporter A1 (ABCA1), the defective molecule in Tangier disease, has been shown to stimulate phospholipid and cholesterol efflux to apolipoprotein A-I (apoA-I); however, little is known concerning the cellular cholesterol pools that act as the source of cholesterol for ABCA1-mediated efflux. We observed a higher level of isotopic and mass cholesterol efflux from mouse peritoneal macrophages labeled with [(3)H]cholesterol/acetyl low density lipoprotein (where cholesterol accumulates in late endosomes and lysosomes) compared with cells labeled with [(3)H]cholesterol with 10% fetal bovine serum, suggesting that late endosomes/lysosomes act as a preferential source of cholesterol for ABCA1-mediated efflux. Consistent with this idea, macrophages from Niemann-Pick C1 mice that have an inability to exit cholesterol from late endosomes/lysosomes showed a profound defect in cholesterol efflux to apoA-I. In contrast, phospholipid efflux to apoA-I was normal in Niemann-Pick C1 macrophages, as was cholesterol efflux following plasma membrane cholesterol labeling. These results suggest that cholesterol deposited in late endosomes/lysosomes preferentially acts as a source of cholesterol for ABCA1-mediated cholesterol efflux.     

Cholesterol and plant sterol efflux from cultured intestinal epithelial cells is mediated by ATP-binding cassette transporters

In this study we analyzed functions of ATP-binding cassette (ABC) transporters involved in sterol transport from Caco-2 cells. Treatment with a synthetic liver x receptor ligand elevated both mRNA and protein levels of ABCG5, G8, and ABCA1. The ligand stimulated cholesterol efflux, suggesting that ABC transporters are involved in it. To identify the acceptors of cholesterol, potential molecules such as apolipoprotein A-I, glycocholic acid, phosphatidylcholine, and bile acid micelles were added to the medium. Apo A-I, a known acceptor of cholesterol transported by ABCA1, elevated cholesterol efflux on the basal side, whereas the others raised cholesterol efflux on the apical side. Moreover, bile acid micelles preferentially augmented plant sterol efflux rather than cholesterol. Finally, in HEK293 cells stably expressing ABCG5/G8, bile acid micelle-mediated sterol efflux was significantly accelerated. These results indicate that ABCG5/G8, unlike ABCA1, together with bile acids should participate in sterol efflux on the apical surface of Caco-2 cells.     

The role of ATP-binding cassette transporters in bacterial pathogenicity

The ATP-binding cassette transporter superfamily is present in all three domains of life. This ubiquitous class of integral membrane proteins have diverse biological functions, but their fundamental role involves the unidirectional translocation of compounds across cellular membranes in an ATP coupled process. The importance of this class of proteins in eukaryotic systems is well established as typified by their association with genetic diseases and roles in the multi-drug resistance of cancer. In stark contrast, the ABC transporters of prokaryotes have not been exhaustively investigated due to the sheer number of different roles and organisms in which they function. In this review, we examine the breadth of functions associated with microbial ABC transporters in the context of their contribution to bacterial pathogenicity and virulence.     

结核分枝杆菌ABC转运蛋白与物质的跨膜转运

结核分枝杆菌作为一种胞内寄生菌,主要存在于巨噬细胞吞噬体内,并且通过与宿主细胞竞争摄取营养物质、主动排出有毒物质来维持生存。因此,参与上述过程的ABC转运蛋白在结核分枝杆菌的致病中发挥着举足轻重的作用。已有报道结核分枝杆菌基因组编码了38个ABC转运蛋白。这类蛋白质有着广泛的底物结合谱,参与了无机离子、糖类、氨基酸、寡肽、药物等多种物质的跨膜转运。本文将对结核分枝杆菌编码的ABC转运蛋白超家族中的不同成员及其底物特异性、转运机制以及与毒力的关系的研究进展进行综述。     

Review. Structure and mechanism of ATP-binding cassette transporters

ATP-binding cassette (ABC) transporters constitute a large superfamily of integral membrane proteins that includes both importers and exporters. In recent years, several structures of complete ABC transporters have been determined by X-ray crystallography. These structures suggest a mechanism by which binding and hydrolysis of ATP by the cytoplasmic, nucleotide-binding domains control the conformation of the transmembrane domains and therefore which side of the membrane the translocation pathway is exposed to. A basic, conserved two-state mechanism can explain active transport of both ABC importers and ABC exporters, but various questions remain unresolved. In this article, I will review some of the crystal structures and the mechanistic insight gained from them. Future challenges for a better understanding of the mechanism of ABC transporters will be outlined.     

The correlation of ATP-binding cassette 1 mRNA levels with cholesterol efflux from various cell lines

Studies show that lipid-free apoA-I stimulates release of cholesterol and phospholipid from fibroblasts and macrophages. ATP-binding cassette 1 (ABC1) is implicated in this release and has been identified as the genetic defect in Tangier disease, evidence that ABC1 is critical to the biogenesis of high density lipoprotein. We quantified levels of ABC1 mRNA, protein, and cholesterol efflux from J774 mouse macrophages +/- exposure to a cAMP analog. Up-regulating ABC1 mRNA correlated to increased cholesterol efflux in a dose- and time-dependent manner. mRNA levels rose after 15 min of exposure while protein levels rose after 1 h, with increased efflux 2-4 h post-treatment. In contrast to cells from wild-type mice, peritoneal macrophages from the Abc1 -/- mouse showed a lower level of basal efflux and no increase with cAMP treatment. The stimulation of efflux exhibits specificity for apoA-I, high density lipoprotein, and other apolipoproteins as cholesterol acceptors, but not for small unilamellar vesicles, bile acid micelles, or cyclodextrin. We have studied a number of cell types and found that while other cell lines express ABC1 constitutively, only J774 and elicited mouse macrophages show a substantial increase of mRNA and efflux with cAMP treatment. ApoA-I-stimulated efflux was detected from the majority of cell lines examined, independent of treatment.     

ATP-binding cassette transporter A1 (ABCA1) functions as a cholesterol efflux regulatory protein

ABCA1, an ATP-binding cassette transporter mutated in Tangier disease, promotes cellular phospholipid and cholesterol efflux by loading free apoA-I with these lipids. This process involves binding of apoA-I to the cell surface and phospholipid translocation by ABCA1. The goals of this study were to examine the relationship between ABCA1-mediated lipid efflux and apolipoprotein binding and to determine whether phospholipid and cholesterol efflux are coupled. Inhibition of lipid efflux by glybenclamide treatment or by mutation of the ATP-binding cassette of ABCA1 showed a close correlation between lipid efflux, the binding of apoA-I to cells, and cross-linking of apoA-I to ABCA1. The data suggest that a functionally important apoA-I binding site exists on ABCA1 and that the binding site could also involve lipids. After using cyclodextrin preincubation to deplete cellular cholesterol, ABCA1-mediated cholesterol efflux was abolished but phospholipid efflux and the binding of apoA-I were unaffected. The conditioned media from cyclodextrin-pretreated, ABCA1-expressing cells readily promoted cholesterol efflux when added to fresh cells not expressing ABCA1, indicating that cholesterol efflux can be dissociated from phospholipid efflux. Further, using a photoactivatable cholesterol analog, we showed that ABCA1 did not bind cholesterol directly, even though several other cholesterol-binding proteins specifically bound the cholesterol analog. The data suggest that the binding of apoA-I to ABCA1 leads to the formation of phospholipid-apoA-I complexes, which subsequently promote cholesterol efflux in an autocrine or paracrine fashion.     

ATP-binding cassette transporters modulate both coelenterazine- and D-luciferin-based bioluminescence imaging

Bioluminescence imaging (BLI) of luciferase reporters provides a cost-effective and sensitive means to image biological processes. However, transport of luciferase substrates across the cell membrane does affect BLI readout intensity from intact living cells. To investigate the effect of ATP-binding cassette (ABC) transporters on BLI readout, we generated click beetle (cLuc), firefly (fLuc), Renilla (rLuc), and Gaussia (gLuc) luciferase HEK-293 reporter cells that overexpressed different ABC transporters (ABCB1, ABCC1, and ABCG2). In vitro studies showed a significant BLI intensity decrease in intact cells compared to cell lysates, when ABCG2 was overexpressed in HEK-293/cLuc, fLuc, and rLuc cells. Selective ABC transporter inhibitors were also applied. Inhibition of ABCG2 activity increased the BLI intensity more than two-fold in HEK-293/cLuc, fLuc, and rLuc cells; inhibition of ABCB1 elevated the BLI intensity two-fold only in HEK-293/rLuc cells. BLI of xenografts derived from HEK-293/ABC transporter/luciferase reporter cells confirmed the results of inhibitor treatment in vivo. These findings demonstrate that coelenterazine-based rLuc-BLI intensity can be modulated by ABCB1 and ABCG2. ABCG2 modulates d-luciferin-based BLI in a luciferase type-independent manner. Little ABC transporter effect on gLuc-BLI intensity is observed because a large fraction of gLuc is secreted. The expression level of ABC transporters is one key factor affecting BLI intensity, and this may be particularly important in luciferase-based applications in stem cell research.     

节肢动物ABC转运蛋白及其介导的杀虫剂抗性

腺苷三磷酸结合盒转运蛋白（ATP-binding cassette transporter),简称ABC转运蛋白（ABC transporter）,是继细胞色素P450单加氧酶、羧酸酯酶、谷胱甘肽S-转移酶之后又一类参与解毒作用的重要蛋白家族,因其在杀虫剂解毒等方面起着非常重要的作用,近年来逐渐受到广泛关注。ABC转运蛋白是一大类跨膜蛋白,其核心结构通常由4个结构域组成,包括2个高度疏水的跨膜结构域（transmembrane domains , TMD）和2个核苷酸结合域（nucleotide binding domains, NBD）。根据序列相似性和保守结构域,可以把ABC转运蛋白家族分为8个亚家族,每个亚家族的成员数及功能不同。这类蛋白在各种生物体内均有分布,其主要功能包括转运物质、信号传导、细胞表面受体及参与细胞内DNA修复,转录及调节基因的表达过程等。此外,近年来的研究表明,ABC转运蛋白的突变或过表达不仅与节肢动物对化学农药的抗药性密切相关,而且在抗Bt毒素方面也起着非常重要的作用,对转Bt作物造成严重威胁。本文综述了节肢动物ABC转运蛋白的结构,ATP水解介导的作用机制,亚家族的分类、结构及生理功能,以及由ABC转运蛋白介导的抗药性研究进展,旨在深入了解ABC转运蛋白的研究现状及其在节肢动物抗药性方面的作用,为阐明节肢动物抗药性机制提供新的理论依据,对改进农业害虫的抗性监测和治理策略也具有一定的指导意义。     

ATP-binding cassette (ABC) transporters in normal and pathological lung

ATP-binding cassette (ABC) transporters are a family of transmembrane proteins that can transport a wide variety of substrates across biological membranes in an energy-dependent manner. Many ABC transporters such as P-glycoprotein (P-gp), multidrug resistance-associated protein 1 (MRP1) and breast cancer resistance protein (BCRP) are highly expressed in bronchial epithelium. This review aims to give new insights in the possible functions of ABC molecules in the lung in view of their expression in different cell types. Furthermore, their role in protection against noxious compounds, e.g. air pollutants and cigarette smoke components, will be discussed as well as the (mal)function in normal and pathological lung. Several pulmonary drugs are substrates for ABC transporters and therefore, the delivery of these drugs to the site of action may be highly dependent on the presence and activity of many ABC transporters in several cell types. Three ABC transporters are known to play an important role in lung functioning. Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene can cause cystic fibrosis, and mutations in ABCA1 and ABCA3 are responsible for respectively Tangier disease and fatal surfactant deficiency. The role of altered function of ABC transporters in highly prevalent pulmonary diseases such as asthma or chronic obstructive pulmonary disease (COPD) have hardly been investigated so far. We especially focused on polymorphisms, knock-out mice models and in vitro results of pulmonary research. Insight in the function of ABC transporters in the lung may open new ways to facilitate treatment of lung diseases.     

Photobiomodulation therapy promotes the ATP-binding cassette transporter A1-dependent cholesterol efflux in macrophage to ameliorate atherosclerosis

Atherosclerosis is a chronic inflammatory disease related to a massive accumulation of cholesterol in the artery wall. Photobiomodulation therapy (PBMT) has been reported to possess cardioprotective effects but has no consensus on the underlying mechanisms. Here, we aimed to investigate whether PBMT could ameliorate atherosclerosis and explore the potential molecular mechanisms. The Apolipoprotein E (ApoE)^−/− mice were fed with western diet (WD) for 18 weeks and treated with PBMT once a day in the last 10 weeks. Quantification based on Oil red O-stained aortas showed that the average plaque area decreased 8.306 ± 2.012% after PBMT (P < .05). Meanwhile, we observed that high-density lipoprotein cholesterol level in WD + PBMT mice increased from 0.309 ± 0.037 to 0.472 ± 0.038 nmol/L (P < .05) compared with WD mice. The further results suggested that PBMT could promote cholesterol efflux from lipid-loaded primary peritoneal macrophages and inhibit foam cells formation via up-regulating the ATP-binding cassette transporters A1 expression. A contributing mechanism involved in activating the phosphatidylinositol 3-kinases/protein kinase C zeta/specificity protein 1 signalling cascade. Our study outlines that PBMT has a protective role on atherosclerosis by promoting macrophages cholesterol efflux and provides a new strategy for treating atherosclerosis.