ABC转运蛋白研究的新进展

ABC转运蛋白主要包括P-糖蛋白、多药耐药相关蛋白和乳腺癌耐药蛋白,它们属于同一家族,具有保守的功能结构域和多样化的生物学功能。ABC转运蛋白部分成员的过表达与肿瘤细胞的多药耐药性（MDR）密切相关,是导致化疗失败的主要原因。随着对MDR机制认识的深入,针对多药耐药蛋白的特异结构域已设计出多种形式的MDR逆转药物。近年来发现,ABC转运蛋白广泛存在于多种正常的组织和器官,参与药物和内、外源毒素的吸收、分布和排泄,行使解毒和防御保护的作用。因此,通过转植ABC转运蛋白基因有可能降低经济鱼类、虾等水产品中有毒污染物的积累。     

ABC细胞膜转运蛋白及其介导的细胞多药耐药研究进展

ABC细胞膜转运蛋白是一个能转运多种底物的蛋白质家族,其在宿主对异物的防御机制和肿瘤细胞对抗癌药物的耐药性中发挥重要作用。ABC转运蛋白能将已进人细胞的外源性物质从胞内泵出胞外,是造成肿瘤细胞多药耐药的主要原因,其基因表达水平与细胞内药物浓度和耐药程度密切相关。近年来,肿瘤细胞多药耐药性研究炙手可热。我们简要综述ABC细胞膜转运蛋白的特点、分布、表达及其介导的细胞多药耐药方面的研究进展。     

多药耐药相关蛋白1及其与病原体感染和耐药的相关性

多药耐药相关蛋白1(multidrug resistance-associated protein 1,MRP1)是ATP结合盒转运蛋白超家族的ABCC亚家族成员,在人体内分布广泛,转运底物众多,具有重要的生理学、病理学和药理学功能。除与肿瘤及抗肿瘤药物耐药、炎症性和免疫性疾病、心血管疾病、神经系统疾病等密切相关外,近年来MRP1在感染性疾病中的作用也日益受到关注。该文就MRP1的结构、功能和调控及其与病原体感染和耐药相关性的研究进展作一综述。     

MRP基因与肿瘤的多药耐药性

在人肿瘤非典型性多药耐药机制的研究中发现了一个新的基因——多药耐药相关蛋白基因（MRP）.该基因位于人16号染色体P13∶3,编码1 531个氨基酸.其产物为多药耐药相关蛋白（MRP）,分子质量190 ku,故又名p190. MRP属ABC超家族成员,主要分布在细胞的质膜上.MRP的功能可能是在能量依赖的外排系统中发挥作用.除了一些肿瘤细胞系外,MRP基因的高表达还见于一些血液系肿瘤及乳腺癌等.MRP基因的高表达还可能与某些肿瘤的复发和预后有关.     

乳腺癌耐药蛋白的研究进展

乳腺癌耐药相关蛋白(BCRP/ABCG2)是新近发现的ATP结合盒(Adenosine triphosphate-binding cassette,ABC)膜转运蛋白超家族成员。它作为细胞膜上的药物排出泵,可以将一系列细胞毒药物转运至胞外从而介导肿瘤细胞多药耐药。在很多血液肿瘤和实体瘤中均检测到ABCG2表达。ABCG2在肿瘤的多药耐药上发挥重要作用。本文对ABCG2的发现、基因表达特征、与造血干细胞分化的关系、转运底物及其耐药逆转和临床意义等方面的研究进展进行综述。     

ABC转运蛋白结构及在植物病原真菌中的功能研究进展

ABC(ATP-binding cassette)转运蛋白是最大的膜转运蛋白超家族之一,其主要功能是利用ATP水解产生的能量将底物进行逆浓度梯度运输.所有生物体都含有大量ABC蛋白.ABC蛋白位于细胞的不同空间,如细胞膜、液泡、线粒体和过氧化物酶体.通常,ABC转运蛋白由跨膜结构域(TMD)和核苷酸结合结构域(NBD)组成,分别与底物和ATP结合.NBD执行与ATP结合和水解,是ABC转运蛋白的动力引擎,TMD识别特异性配体.大多数ABC转运蛋白最初是通过研究生物体耐药性而被发现的,包括多效耐药(PDR)和多药耐药(MDR).本文对ABC转运蛋白的结构及作用机制,以及植物病原真菌中ABC转运蛋白功能的研究进展进行综述.     

P-糖蛋白对结肠癌多药耐药的影响

结肠癌是常见的消化道恶性肿瘤。对术后患者以及无法采用手术治疗的患者,临床多采用化疗、放疗等综合性治疗方法。随着大量化疗药物在临床的广泛使用,结肠癌多药耐药性成为化疗失败的最主要原因。研究表明,P-糖蛋白(P-glycoprotein, P-gp)作为ATP结合盒(ABC)转运蛋白超家族成员之一,与多种肿瘤的多药耐药相关,其介导的多药耐药已经成为目前研究的热点。本文旨在通过对P-糖蛋白的结构、耐药机制以及逆转P-糖蛋白介导的结肠癌多药耐药新发现进行阐述,引导读者对P-糖蛋白在结肠癌多药耐药中的作用有更深入的了解。     

肿瘤多药耐药性中的ABC转运蛋白及其调节

ABC转运蛋白是一类利用ATP水解能量,逆浓度方向将一系列化合物转运通过膜结构的膜蛋白,这一类蛋白能转运离子,糖,氨基酸,维生素,多肽,多糖,激素,脂类及生物异源物质.P-糖蛋白(P-gp)、多药耐药相关蛋白(MRP)和乳腺癌耐药蛋白(BCRP)等ABC转运蛋白还具有转运抗癌药物的能力,因此对化疗的有效性有负面的影响.近年来,许多研究涉及到如何逆转由ABC转 运蛋白引起的肿瘤多药耐药性.本文概述了近年来在蛋白水平,mRNA水平或DNA水平上对ABC转运蛋白调控的研究.     

c-Myc通过调控ATP结合盒转运蛋白表达对CD133~+肠癌干细胞耐药性的影响

本文旨在探讨c-Myc通过调控ATP结合盒(ATP-binding cassette,ABC)转运蛋白表达对CD133~+肠癌干细胞耐药性的影响及机制。以人肠癌HT29细胞系为模型,通过流式细胞分选术获得CD133~+肠癌干细胞,以核心基因c-Myc为靶点设计小干扰RNA(siRNA),靶向抑制CD133~+细胞中c-Myc基因表达;免疫印迹法检测c-Myc沉默后ABC转运蛋白家族成员(ABCG2、MDR-1和ABCB5)表达水平变化;药敏实验检测肠癌干细胞对化疗药物(5-FU或/和奥沙利铂)的耐药性。结果显示,CD133~+细胞高表达核心基因c-Myc,si RNA显著抑制肠癌干细胞中c-Myc的表达水平;CD133~+肠癌干细胞对化疗药物(5-FU或/和奥沙利铂)具有耐药性;si RNA通过抑制c-Myc下调CD133~+肠癌干细胞ABC转运蛋白(ABCG2、MDR-1和ABCB5)的表达水平;靶向抑制c-Myc表达提高CD133~+肠癌干细胞对化疗药物的敏感性(P0.05)。以上结果表明,c-Myc通过调控ABC转运蛋白表达水平参与肿瘤干细胞的耐药机理,靶向抑制c-Myc能提高CD133~+肠癌干细胞对化疗药物的敏感性。     

肺癌细胞微生态成分分析及其在耐药性方面的作用及机制

目的探析肺癌细胞微生态环境中蛋白成分及其与人肺癌多药耐药细胞系SPC-A1/TAX耐药性之间的关系。方法通过基因芯片、蛋白质谱实验并借助S-P法以及MTS法来分析蛋白与耐药之间的关系。结果 SPC-A1/TAX与SPC-A1相比,除了对紫杉醇具有耐受性外,对其他抗肿瘤药物也有一定程度的耐药性。SPC-A1中,GST-π以及MRP的阳性率均为0,而SPC-A1/TAX中两者的阳性率均为100%,对其他抗肿瘤药物也有一定程度的耐药性,耐药倍数分别为6.34、2.42、13.51、2.65、9.80、7.00。结论多药耐药相关蛋白及谷胱甘肽-S-转移酶π与人肺癌多药耐药细胞系SPC-A1/TAX的耐药性密切相关,是导致其具有耐药性的重要机制。     

Mapping of the MRPm5 epitope to the cytosolic region between transmembrane helices 13 and 14 in the drug and organic anion transporter, MRP1 (ABCC1)

Multidrug resistance in human tumour cells is often associated with increased expression of the 190kDa multidrug resistance protein, MRP1, that belongs to the ATP-binding cassette superfamily of transport proteins. MRP1 is also an efficient transporter of many organic anions. In the present study, we have mapped the epitope of the MRP1-specific murine monoclonal antibody (MAb) MRPm5 to the decapeptide (1063)FFERTPSGNL(1072) located in the cytoplasmic loop (CL6) linking transmembrane helices 13 and 14 in the third membrane spanning domain of the protein. Several amino acids in the cytoplasmic loops of MRP1 have been reported to be important for its transport function; nevertheless, MAb MRPm5 does not inhibit vesicular uptake of the high affinity substrate leukotriene C(4). None of the other MRP1-reactive MAbs described to date map to CL6 of MRP1 which in turn enhances the utility of MAb MRPm5 for both clinical and experimental investigations of this transporter.     

Overexpression,purification, and functional characterization of ATP-binding cassette transporters in the yeast,Pichia pastoris

The ATP-binding cassette (ABC) transporter superfamily is a large gene family that has been highly conserved throughout evolution. The physiological importance of these membrane transporters is highlighted by the large variety of substrates they transport, and by the observation that mutations in many of them cause heritable diseases in human. Likewise, overexpression of certain ABC transporters, such as P-glycoprotein and members of the multidrug resistance associated protein (MRP) family, is associated with multidrug resistance in various cells and organisms. Understanding the structure and molecular mechanisms of transport of the ABC transporters in normal tissues and their possibly altered function in human diseases requires large amounts of purified and active proteins. For this, efficient expression systems are needed. The methylotrophic yeast Pichia pastoris has proven to be an efficient and inexpensive experimental model for high-level expression of many proteins, including ABC transporters. In the present review, we will summarize recent advances on the use of this system for the expression, purification, and functional characterization of P-glycoprotein and two members of the MRP subfamily.     

Overexpression, purification, and functional characterization of ATP-binding cassette transporters in the yeast, Pichia pastoris

The ATP-binding cassette (ABC) transporter superfamily is a large gene family that has been highly conserved throughout evolution. The physiological importance of these membrane transporters is highlighted by the large variety of substrates they transport, and by the observation that mutations in many of them cause heritable diseases in human. Likewise, overexpression of certain ABC transporters, such as P-glycoprotein and members of the multidrug resistance associated protein (MRP) family, is associated with multidrug resistance in various cells and organisms. Understanding the structure and molecular mechanisms of transport of the ABC transporters in normal tissues and their possibly altered function in human diseases requires large amounts of purified and active proteins. For this, efficient expression systems are needed. The methylotrophic yeast Pichia pastoris has proven to be an efficient and inexpensive experimental model for high-level expression of many proteins, including ABC transporters. In the present review, we will summarize recent advances on the use of this system for the expression, purification, and functional characterization of P-glycoprotein and two members of the MRP subfamily.     

Structural, mechanistic and clinical aspects of MRP1

The cDNA encoding ATP-binding cassette (ABC) multidrug resistance protein MRP1 was originally cloned from a drug-selected lung cancer cell line resistant to multiple natural product chemotherapeutic agents. MRP1 is the founder of a branch of the ABC superfamily whose members (from species as diverse as plants and yeast to mammals) share several distinguishing structural features that may contribute to functional and mechanistic similarities among this subgroup of transport proteins. In addition to its role in resistance to natural product drugs, MRP1 (and related proteins) functions as a primary active transporter of structurally diverse organic anions, many of which are formed by the biotransformation of various endo- and xenobiotics by Phase II conjugating enzymes, such as the glutathione S-transferases. MRP1 is involved in a number of glutathione-related cellular processes. Glutathione also appears to play a key role in MRP1-mediated drug resistance. This article reviews the discovery of MRP1 and its relationships with other ABC superfamily members, and summarizes current knowledge of the structure, transport functions and relevance of this protein to in vitro and clinical multidrug resistance.     

The Role of Half-Transporters in Multidrug Resistance

ATP-binding cassette proteins comprise a superfamily of transporter proteins, a subset of which have been implicated in multidrug resistance. Although P-glycoprotein was described over 15 years ago, the recent expansion in the number of transporters identified has prompted renewed interest in the role of drug transporters in clinical drug resistance. These newly identified transporters include additional members of the MRP family, ABC2, and a new half-transporter, MXR/BCRP/ABCP1. This half-transporter confers high levels of resistance to mitoxantrone, anthracyclines, and the camptothecins SN-38 and topotecan. At 72 kDa, MXR localizes to the plasma membrane in cells which highly overexpress the protein either through gene amplification or though gene rearrangement. Future studies will be aimed at identifying an inhibitor, and attempting to translate recognition of this new transporter into a target for anticancer treatment.     

Mutation of Trp1254 in the multispecific organic anion transporter, multidrug resistance protein 2 (MRP2) (ABCC2), alters substrate specificity and results in loss of methotrexate transport activity

The ATP-binding cassette (ABC) proteins comprise a large superfamily of transmembrane transporters that utilize the energy of ATP hydrolysis to translocate their substrates across biological membranes. Multidrug resistance protein (MRP) 2 (ABCC2) belongs to subfamily C of the ABC superfamily and, when overexpressed in tumor cells, confers resistance to a wide variety of anticancer chemotherapeutic agents. MRP2 is also an active transporter of organic anions such as methotrexate (MTX), estradiol glucuronide (E217betaG), and leukotriene C4 and is located on the apical membrane of polarized cells including hepatocytes where it acts as a biliary transporter. We recently identified a highly conserved tryptophan residue in the related MRP1 that is critical for the substrate specificity of this protein. In the present study, we have examined the effect of replacing the analogous tryptophan residue at position 1254 of MRP2. We found that only nonconservative substitutions (Ala and Cys) of Trp1254 eliminated [3H]E217betaG transport by MRP2, whereas more conservative substitutions (Phe and Tyr) had no effect. In addition, only the most conservatively substituted mutant (W1254Y) transported [3H]leukotriene C4, whereas all other substitutions eliminated transport of this substrate. On the other hand, all substitutions of Trp1254 eliminated transport of [3H]MTX. Finally, we found that sulfinpyrazone stimulated [3H]E217betaG transport by wild-type MRP2 4-fold, whereas transport by the Trp1254 substituted mutants was enhanced 6-10-fold. In contrast, sulfinpyrazone failed to stimulate [3H]MTX transport by either wild-type MRP2 or the MRP2-Trp1254 mutants. Taken together, our results demonstrate that Trp1254 plays an important role in the ability of MRP2 to transport conjugated organic anions and identify this amino acid in the putative last transmembrane segment (TM17) of this ABC protein as being critical for transport of MTX.     

Structural Insights into Transporter-Mediated Drug Resistance in Infectious Diseases

Infectious diseases present a major threat to public health globally. Pathogens can acquire resistance to anti-infectious agents via several means including transporter-mediated efflux. Typically, multidrug transporters feature spacious, dynamic, and chemically malleable binding sites to aid in the recognition and transport of chemically diverse substrates across cell membranes. Here, we discuss recent structural investigations of multidrug transporters involved in resistance to infectious diseases that belong to the ATP-binding cassette (ABC) superfamily, the major facilitator superfamily (MFS), the drug/metabolite transporter (DMT) superfamily, the multidrug and toxic compound extrusion (MATE) family, the small multidrug resistance (SMR) family, and the resistance-nodulation-division (RND) superfamily. These structural insights provide invaluable information for understanding and combatting multidrug resistance.     

Towards the molecular mechanism of prokaryotic and eukaryotic multidrug transporters.

Due to their ability to extrude structurally dissimilar cytotoxic drugs out of the cell, multidrug transporters are able to reduce the cytoplasmic drug concentration, and, hence, are able to confer drug resistance on human cancer cells and pathogenic microorganisms. This review will focus on the molecular properties of two bacterial multidrug transporters, the ATP-binding cassette transporter LmrA and the proton motive force-dependent major facilitator superfamily transporter LmrP, which each represent a major class of multidrug transport proteins encountered in pro- and eukaryotic cells. In spite of the structural differences between LmrA and LmrP, the molecular bases of their drug transport activity may turn out to be more similar than might currently appear.     

Structure-function analysis of multidrug transporters in Lactococcus lactis

The active extrusion of cytotoxic compounds from the cell by multidrug transporters is one of the major causes of failure of chemotherapeutic treatment of tumor cells and of infections by pathogenic microorganisms. A multidrug transporter in Lactococcus lactis, LmrA, is a member of the ATP-binding cassette (ABC) superfamily and a bacterial homolog of the human multidrug resistance P-glycoprotein. Another multidrug transporter in L. lactis, LmrP, belongs to the major facilitator superfamily, and is one example of a rapidly expanding group of secondary multidrug transporters in microorganisms. Thus, LmrA and LmrP are transport proteins with very different protein structures, which use different mechanisms of energy coupling to transport drugs out of the cell. Surprisingly, both proteins have overlapping specificities for drugs, are inhibited by the same set of modulators, and transport drugs via a similar transport mechanism. The structure-function relationships that dictate drug recognition and transport by LmrP and LmrA represent an intriguing area of research.