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Sample records for abc atp-binding cassette

  1. Human ATP-binding cassette (ABC transporter family

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    Vasiliou Vasilis

    2009-04-01

    Full Text Available Abstract There exist four fundamentally different classes of membrane-bound transport proteins: ion channels; transporters; aquaporins; and ATP-powered pumps. ATP-binding cassette (ABC transporters are an example of ATP-dependent pumps. ABC transporters are ubiquitous membrane-bound proteins, present in all prokaryotes, as well as plants, fungi, yeast and animals. These pumps can move substrates in (influx or out (efflux of cells. In mammals, ABC transporters are expressed predominantly in the liver, intestine, blood-brain barrier, blood-testis barrier, placenta and kidney. ABC proteins transport a number of endogenous substrates, including inorganic anions, metal ions, peptides, amino acids, sugars and a large number of hydrophobic compounds and metabolites across the plasma membrane, and also across intracellular membranes. The human genome contains 49 ABC genes, arranged in eight subfamilies and named via divergent evolution. That ABC genes are important is underscored by the fact that mutations in at least I I of these genes are already known to cause severe inherited diseases (eg cystic fibrosis and X-linked adrenoleukodystrophy [X-ALD]. ABC transporters also participate in the movement of most drugs and their metabolites across cell surface and cellular organelle membranes; thus, defects in these genes can be important in terms of cancer therapy, pharmacokinetics and innumerable pharmacogenetic disorders.

  2. Evolutionary relationships of ATP-Binding Cassette (ABC) uptake porters.

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    Zheng, Wei Hao; Västermark, Åke; Shlykov, Maksim A; Reddy, Vamsee; Sun, Eric I; Saier, Milton H

    2013-05-06

    The ATP-Binding Cassette (ABC) functional superfamily includes integral transmembrane exporters that have evolved three times independently, forming three families termed ABC1, ABC2 and ABC3, upon which monophyletic ATPases have been superimposed for energy-coupling purposes [e.g., J Membr Biol 231(1):1-10, 2009]. The goal of the work reported in this communication was to understand how the integral membrane constituents of ABC uptake transporters with different numbers of predicted or established transmembrane segments (TMSs) evolved. In a few cases, high resolution 3-dimensional structures were available, and in these cases, their structures plus primary sequence analyses allowed us to predict evolutionary pathways of origin. All of the 35 currently recognized families of ABC uptake proteins except for one (family 21) were shown to be homologous using quantitative statistical methods. These methods involved using established programs that compare native protein sequences with each other, after having compared each sequence with thousands of its own shuffled sequences, to gain evidence for homology. Topological analyses suggested that these porters contain numbers of TMSs ranging from four or five to twenty. Intragenic duplication events occurred multiple times during the evolution of these porters. They originated from a simple primordial protein containing 3 TMSs which duplicated to 6 TMSs, and then produced porters of the various topologies via insertions, deletions and further duplications. Except for family 21 which proved to be related to ABC1 exporters, they are all related to members of the previously identified ABC2 exporter family. Duplications that occurred in addition to the primordial 3 → 6 duplication included 5 → 10, 6 → 12 and 10 → 20 TMSs. In one case, protein topologies were uncertain as different programs gave discrepant predictions. It could not be concluded with certainty whether a 4 TMS ancestral protein or a 5 TMS ancestral protein

  3. Inventory and comparative analysis of rice and Arabidopsis ATP-binding cassette (ABC) systems.

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    Garcia, Olivier; Bouige, Philippe; Forestier, Cyrille; Dassa, Elie

    2004-10-08

    ATP-binding cassette (ABC) proteins constitute a large superfamily found in all kingdoms of living organisms. The recent completion of two draft sequences of the rice (Oryza sativa) genome allowed us to analyze and classify its ABC proteins and to compare to those in Arabidopsis thaliana. We identified a similar number of ABC proteins in rice and Arabidopsis (121 versus 120), despite the rice genome being more than three times the size of Arabidopsis. Both Arabidopsis and rice have representative members in all seven major subfamilies of ABC ATPases (A to G) commonly found in eukaryotes. This comparative analysis allowed the detection of 29 potential orthologous sequences in Arabidopsis and rice. However, plant share with prokaryotes a specific set of ABC systems that is not detected in animals. These ABC systems might be inherited from the cyanobacterial ancestor of chloroplasts. The present work provides the first complete inventory of rice ABC proteins and an updated inventory of those proteins in Arabidopsis.

  4. ATP-binding cassette (ABC) proteins in aquatic invertebrates: Evolutionary significance and application in marine ecotoxicology.

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    Jeong, Chang-Bum; Kim, Hui-Su; Kang, Hye-Min; Lee, Jae-Seong

    2017-04-01

    The ATP-binding cassette (ABC) protein superfamily is known to play a fundamental role in biological processes and is highly conserved across animal taxa. The ABC proteins function as active transporters for multiple substrates across the cellular membrane by ATP hydrolysis. As this superfamily is derived from a common ancestor, ABC genes have evolved via lineage-specific duplications through the process of adaptation. In this review, we summarized information about the ABC gene families in aquatic invertebrates, considering their evolution and putative functions in defense mechanisms. Phylogenetic analysis was conducted to examine the evolutionary significance of ABC gene families in aquatic invertebrates. Particularly, a massive expansion of multixenobiotic resistance (MXR)-mediated efflux transporters was identified in the absence of the ABCG2 (BCRP) gene in Ecdysozoa and Platyzoa, suggesting that a loss of Abcg2 gene occurred sporadically in these species during divergence of Protostome to Lophotrochozoa. Furthermore, in aquatic invertebrates, the ecotoxicological significance of MXR is discussed while considering the role of MXR-mediated efflux transporters in response to various environmental pollutants. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. ATP binding cassette (ABC) transporters: expression and clinical value in glioblastoma.

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    Dréan, Antonin; Rosenberg, Shai; Lejeune, François-Xavier; Goli, Larissa; Nadaradjane, Aravindan Arun; Guehennec, Jérémy; Schmitt, Charlotte; Verreault, Maïté; Bielle, Franck; Mokhtari, Karima; Sanson, Marc; Carpentier, Alexandre; Delattre, Jean-Yves; Idbaih, Ahmed

    2018-03-08

    ATP-binding cassette transporters (ABC transporters) regulate traffic of multiple compounds, including chemotherapeutic agents, through biological membranes. They are expressed by multiple cell types and have been implicated in the drug resistance of some cancer cells. Despite significant research in ABC transporters in the context of many diseases, little is known about their expression and clinical value in glioblastoma (GBM). We analyzed expression of 49 ABC transporters in both commercial and patient-derived GBM cell lines as well as from 51 human GBM tumor biopsies. Using The Cancer Genome Atlas (TCGA) cohort as a training dataset and our cohort as a validation dataset, we also investigated the prognostic value of these ABC transporters in newly diagnosed GBM patients, treated with the standard of care. In contrast to commercial GBM cell lines, GBM-patient derived cell lines (PDCL), grown as neurospheres in a serum-free medium, express ABC transporters similarly to parental tumors. Serum appeared to slightly increase resistance to temozolomide correlating with a tendency for an increased expression of ABCB1. Some differences were observed mainly due to expression of ABC transporters by microenvironmental cells. Together, our data suggest that the efficacy of chemotherapeutic agents may be misestimated in vitro if they are the targets of efflux pumps whose expression can be modulated by serum. Interestingly, several ABC transporters have prognostic value in the TCGA dataset. In our cohort of 51 GBM patients treated with radiation therapy with concurrent and adjuvant temozolomide, ABCA13 overexpression is associated with a decreased progression free survival in univariate (p ABC transporters is: (i) detected in GBM and microenvironmental cells and (ii) better reproduced in GBM-PDCL. ABCA13 expression is an independent prognostic factor in newly diagnosed GBM patients. Further prospective studies are warranted to investigate whether ABCA13 expression can be

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

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    Timmer-Bosscha Hetty

    2005-06-01

    Full Text Available Abstract 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 (malfunction 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.

  7. The role of ATP-binding cassette (ABC) transporters in pathogenesis and multidrug resistance of the wheat pathogen Mycosphaerella graminicola

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    Stergiopoulos, I.

    2003-01-01

    ATP-binding cassette (ABC) transporters are membrane proteins that utilise the energy derived from the hydrolysis of ATP to drive the transport of compounds over biological membranes. They are members of one of the largest protein families to date, present in both pro- and eukaryotic

  8. Predictive Structure and Topology of Peroxisomal ATP-Binding Cassette (ABC) Transporters

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    Andreoletti, Pierre; Raas, Quentin; Gondcaille, Catherine; Cherkaoui-Malki, Mustapha; Trompier, Doriane; Savary, Stéphane

    2017-01-01

    The peroxisomal ATP-binding Cassette (ABC) transporters, which are called ABCD1, ABCD2 and ABCD3, are transmembrane proteins involved in the transport of various lipids that allow their degradation inside the organelle. Defective ABCD1 leads to the accumulation of very long-chain fatty acids and is associated with a complex and severe neurodegenerative disorder called X-linked adrenoleukodystrophy (X-ALD). Although the nucleotide-binding domain is highly conserved and characterized within the ABC transporters family, solid data are missing for the transmembrane domain (TMD) of ABCD proteins. The lack of a clear consensus on the secondary and tertiary structure of the TMDs weakens any structure-function hypothesis based on the very diverse ABCD1 mutations found in X-ALD patients. Therefore, we first reinvestigated thoroughly the structure-function data available and performed refined alignments of ABCD protein sequences. Based on the 2.85  Å resolution crystal structure of the mitochondrial ABC transporter ABCB10, here we propose a structural model of peroxisomal ABCD proteins that specifies the position of the transmembrane and coupling helices, and highlight functional motifs and putative important amino acid residues. PMID:28737695

  9. Predictive Structure and Topology of Peroxisomal ATP-Binding Cassette (ABC Transporters

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    Pierre Andreoletti

    2017-07-01

    Full Text Available The peroxisomal ATP-binding Cassette (ABC transporters, which are called ABCD1, ABCD2 and ABCD3, are transmembrane proteins involved in the transport of various lipids that allow their degradation inside the organelle. Defective ABCD1 leads to the accumulation of very long-chain fatty acids and is associated with a complex and severe neurodegenerative disorder called X-linked adrenoleukodystrophy (X-ALD. Although the nucleotide-binding domain is highly conserved and characterized within the ABC transporters family, solid data are missing for the transmembrane domain (TMD of ABCD proteins. The lack of a clear consensus on the secondary and tertiary structure of the TMDs weakens any structure-function hypothesis based on the very diverse ABCD1 mutations found in X-ALD patients. Therefore, we first reinvestigated thoroughly the structure-function data available and performed refined alignments of ABCD protein sequences. Based on the 2.85  Å resolution crystal structure of the mitochondrial ABC transporter ABCB10, here we propose a structural model of peroxisomal ABCD proteins that specifies the position of the transmembrane and coupling helices, and highlight functional motifs and putative important amino acid residues.

  10. ATP-binding cassette (ABC) transporter expression and localization in sea urchin development.

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    Shipp, Lauren E; Hamdoun, Amro

    2012-06-01

    ATP-binding cassette (ABC) transporters are membrane proteins that regulate intracellular concentrations of myriad compounds and ions. There are >100 ABC transporter predictions in the Strongylocentrotus purpuratus genome, including 40 annotated ABCB, ABCC, and ABCG "multidrug efflux" transporters. Despite the importance of multidrug transporters for protection and signaling, their expression patterns have not been characterized in deuterostome embryos. Sea urchin embryos expressed 20 ABCB, ABCC, and ABCG transporter genes in the first 58 hr of development, from unfertilized egg to early prism. We quantified transcripts of ABCB1a, ABCB4a, ABCC1, ABCC5a, ABCC9a, and ABCG2b, and found that ABCB1a mRNA was 10-100 times more abundant than other transporter mRNAs. In situ hybridization showed ABCB1a was expressed ubiquitously in embryos, while ABCC5a was restricted to secondary mesenchyme cells and their precursors. Fluorescent protein fusions showed localization of ABCB1a on apical cell surfaces, and ABCC5a on basolateral surfaces. Embryos use many ABC transporters with predicted functions in cell signaling, lysosomal and mitochondrial homeostasis, potassium channel regulation, pigmentation, and xenobiotic efflux. Detailed characterization of ABCB1a and ABCC5a revealed that they have different temporal and spatial gene expression profiles and protein localization patterns that correlate to their predicted functions in protection and development, respectively. Copyright © 2012 Wiley Periodicals, Inc.

  11. Inventory and analysis of ATP-binding cassette (ABC) systems in Brugia malayi.

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    Ardelli, B F; Stitt, L E; Tompkins, J B

    2010-07-01

    ABC systems are one of the largest described protein superfamilies. These systems have a domain organization that may contain 1 or more transmembrane domains (ABC_TM1F) and 1 or 2 ATP-binding domains (ABC_2). The functions (e.g., import, export and DNA repair) of these proteins distinguish the 3 classes of ABC systems. Mining and PCR-based cloning were used to identify 33 putative ABC systems from the Brugia malayi genome. There were 31 class 2 genes, commonly called ABC transporters, and 2 class 3 genes. The ABC transporters were divided into subfamilies. Three belonged to subfamily A, 16 to subfamily B, 5 to subfamily C, 1 to subfamily E and 3 to subfamilies F and G, respectively. None were placed in subfamilies D and H. Similar to other ABC systems, the ABC_2 domain of B. malayi genes was conserved and contained the Walker A and B motifs, the signature sequence/linker region and the switch region with the conserved histidine. The ABC_TM1F domain was less conserved. The relative abundance of ABC systems was quantified using real-time reverse transcription PCR and was significantly higher in female adults of B. malayi than in males and microfilaria, particularly those in subfamilies B and C, which are associated with drug resistance.

  12. Inventory and general analysis of the ATP-binding cassette (ABC) gene superfamily in maize (Zea mays L.).

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    Pang, Kaiyuan; Li, Yanjiao; Liu, Menghan; Meng, Zhaodong; Yu, Yanli

    2013-09-10

    The metabolic functions of ATP-binding cassette (or ABC) proteins, one of the largest families of proteins presented in all organisms, have been investigated in many protozoan, animal and plant species. To facilitate more systematic and complicated studies on maize ABC proteins in the future, we present the first complete inventory of these proteins, including 130 open reading frames (ORFs), and provide general descriptions of their classifications, basic structures, typical functions, evolution track analysis and expression profiles. The 130 ORFs were assigned to eight subfamilies based on their structures and homological features. Five of these subfamilies consist of 109 proteins, containing transmembrane domains (TM) performing as transporters. The rest three subfamilies contain 21 soluble proteins involved in various functions other than molecular transport. A comparison of ABC proteins among nine selected species revealed either convergence or divergence in each of the ABC subfamilies. Generally, plant genomes contain far more ABC genes than animal genomes. The expression profiles and evolution track of each maize ABC gene were further investigated, the results of which could provide clues for analyzing their functions. Quantitative real-time polymerase chain reaction experiments (PCR) were conducted to detect induced expression in select ABC genes under several common stresses. This investigation provides valuable information for future research on stress tolerance in plants and potential strategies for enhancing maize production under stressful conditions. Copyright © 2013 Elsevier B.V. All rights reserved.

  13. Genome-wide identification, characterization and phylogenetic analysis of 50 catfish ATP-binding cassette (ABC transporter genes.

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    Shikai Liu

    Full Text Available Although a large set of full-length transcripts was recently assembled in catfish, annotation of large gene families, especially those with duplications, is still a great challenge. Most often, complexities in annotation cause mis-identification and thereby much confusion in the scientific literature. As such, detailed phylogenetic analysis and/or orthology analysis are required for annotation of genes involved in gene families. The ATP-binding cassette (ABC transporter gene superfamily is a large gene family that encodes membrane proteins that transport a diverse set of substrates across membranes, playing important roles in protecting organisms from diverse environment.In this work, we identified a set of 50 ABC transporters in catfish genome. Phylogenetic analysis allowed their identification and annotation into seven subfamilies, including 9 ABCA genes, 12 ABCB genes, 12 ABCC genes, 5 ABCD genes, 2 ABCE genes, 4 ABCF genes and 6 ABCG genes. Most ABC transporters are conserved among vertebrates, though cases of recent gene duplications and gene losses do exist. Gene duplications in catfish were found for ABCA1, ABCB3, ABCB6, ABCC5, ABCD3, ABCE1, ABCF2 and ABCG2.The whole set of catfish ABC transporters provide the essential genomic resources for future biochemical, toxicological and physiological studies of ABC drug efflux transporters. The establishment of orthologies should allow functional inferences with the information from model species, though the function of lineage-specific genes can be distinct because of specific living environment with different selection pressure.

  14. A Survey of the ATP-Binding Cassette (ABC Gene Superfamily in the Salmon Louse (Lepeophtheirus salmonis.

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    Greta Carmona-Antoñanzas

    Full Text Available Salmon lice, Lepeophtheirus salmonis (Krøyer, 1837, are fish ectoparasites causing significant economic damage in the mariculture of Atlantic salmon, Salmo salar Linnaeus, 1758. The control of L. salmonis at fish farms relies to a large extent on treatment with anti-parasitic drugs. A problem related to chemical control is the potential for development of resistance, which in L. salmonis is documented for a number of drug classes including organophosphates, pyrethroids and avermectins. The ATP-binding cassette (ABC gene superfamily is found in all biota and includes a range of drug efflux transporters that can confer drug resistance to cancers and pathogens. Furthermore, some ABC transporters are recognised to be involved in conferral of insecticide resistance. While a number of studies have investigated ABC transporters in L. salmonis, no systematic analysis of the ABC gene family exists for this species. This study presents a genome-wide survey of ABC genes in L. salmonis for which, ABC superfamily members were identified through homology searching of the L. salmonis genome. In addition, ABC proteins were identified in a reference transcriptome of the parasite generated by high-throughput RNA sequencing (RNA-seq of a multi-stage RNA library. Searches of both genome and transcriptome allowed the identification of a total of 33 genes / transcripts coding for ABC proteins, of which 3 were represented only in the genome and 4 only in the transcriptome. Eighteen sequences were assigned to ABC subfamilies known to contain drug transporters, i.e. subfamilies B (4 sequences, C (11 and G (2. The results suggest that the ABC gene family of L. salmonis possesses fewer members than recorded for other arthropods. The present survey of the L. salmonis ABC gene superfamily will provide the basis for further research into potential roles of ABC transporters in the toxicity of salmon delousing agents and as potential mechanisms of drug resistance.

  15. A Survey of the ATP-Binding Cassette (ABC) Gene Superfamily in the Salmon Louse (Lepeophtheirus salmonis).

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    Carmona-Antoñanzas, Greta; Carmichael, Stephen N; Heumann, Jan; Taggart, John B; Gharbi, Karim; Bron, James E; Bekaert, Michaël; Sturm, Armin

    2015-01-01

    Salmon lice, Lepeophtheirus salmonis (Krøyer, 1837), are fish ectoparasites causing significant economic damage in the mariculture of Atlantic salmon, Salmo salar Linnaeus, 1758. The control of L. salmonis at fish farms relies to a large extent on treatment with anti-parasitic drugs. A problem related to chemical control is the potential for development of resistance, which in L. salmonis is documented for a number of drug classes including organophosphates, pyrethroids and avermectins. The ATP-binding cassette (ABC) gene superfamily is found in all biota and includes a range of drug efflux transporters that can confer drug resistance to cancers and pathogens. Furthermore, some ABC transporters are recognised to be involved in conferral of insecticide resistance. While a number of studies have investigated ABC transporters in L. salmonis, no systematic analysis of the ABC gene family exists for this species. This study presents a genome-wide survey of ABC genes in L. salmonis for which, ABC superfamily members were identified through homology searching of the L. salmonis genome. In addition, ABC proteins were identified in a reference transcriptome of the parasite generated by high-throughput RNA sequencing (RNA-seq) of a multi-stage RNA library. Searches of both genome and transcriptome allowed the identification of a total of 33 genes / transcripts coding for ABC proteins, of which 3 were represented only in the genome and 4 only in the transcriptome. Eighteen sequences were assigned to ABC subfamilies known to contain drug transporters, i.e. subfamilies B (4 sequences), C (11) and G (2). The results suggest that the ABC gene family of L. salmonis possesses fewer members than recorded for other arthropods. The present survey of the L. salmonis ABC gene superfamily will provide the basis for further research into potential roles of ABC transporters in the toxicity of salmon delousing agents and as potential mechanisms of drug resistance.

  16. ATP-Binding Cassette (ABC) Transporters of the Human Respiratory Tract Pathogen, Moraxella catarrhalis: Role in Virulence.

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    Murphy, Timothy F; Brauer, Aimee L; Johnson, Antoinette; Kirkham, Charmaine

    2016-01-01

    Moraxella catarrhalis is a human respiratory tract pathogen that causes otitis media (middle ear infections) in children and respiratory tract infections in adults with chronic obstructive pulmonary disease. In view of the huge global burden of disease caused by M. catarrhalis, the development of vaccines to prevent these infections and better approaches to treatment have become priorities. In previous work, we used a genome mining approach that identified three substrate binding proteins (SBPs) of ATP-binding cassette (ABC) transporters as promising candidate vaccine antigens. In the present study, we performed a comprehensive assessment of 19 SBPs of 15 ABC transporter systems in the M. catarrhalis genome by engineering knockout mutants and studying their role in assays that assess mechanisms of infection. The capacity of M. catarrhalis to survive and grow in the nutrient-limited and hostile environment of the human respiratory tract, including intracellular growth, account in part for its virulence. The results show that ABC transporters that mediate uptake of peptides, amino acids, cations and anions play important roles in pathogenesis by enabling M. catarrhalis to 1) grow in nutrient-limited conditions, 2) invade and survive in human respiratory epithelial cells and 3) persist in the lungs in a murine pulmonary clearance model. The knockout mutants of SBPs and ABC transporters showed different patterns of activity in the assay systems, supporting the conclusion that different SBPs and ABC transporters function at different stages in the pathogenesis of infection. These results indicate that ABC transporters are nutritional virulence factors, functioning to enable the survival of M catarrhalis in the diverse microenvironments of the respiratory tract. Based on the role of ABC transporters as virulence factors of M. catarrhalis, these molecules represent potential drug targets to eradicate the organism from the human respiratory tract.

  17. Genome-Wide Identification, Characterization and Phylogenetic Analysis of ATP-Binding Cassette (ABC) Transporter Genes in Common Carp (Cyprinus carpio).

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    Liu, Xiang; Li, Shangqi; Peng, Wenzhu; Feng, Shuaisheng; Feng, Jianxin; Mahboob, Shahid; Al-Ghanim, Khalid A; Xu, Peng

    2016-01-01

    The ATP-binding cassette (ABC) gene family is considered to be one of the largest gene families in all forms of prokaryotic and eukaryotic life. Although the ABC transporter genes have been annotated in some species, detailed information about the ABC superfamily and the evolutionary characterization of ABC genes in common carp (Cyprinus carpio) are still unclear. In this research, we identified 61 ABC transporter genes in the common carp genome. Phylogenetic analysis revealed that they could be classified into seven subfamilies, namely 11 ABCAs, six ABCBs, 19 ABCCs, eight ABCDs, two ABCEs, four ABCFs, and 11 ABCGs. Comparative analysis of the ABC genes in seven vertebrate species including common carp, showed that at least 10 common carp genes were retained from the third round of whole genome duplication, while 12 duplicated ABC genes may have come from the fourth round of whole genome duplication. Gene losses were also observed for 14 ABC genes. Expression profiles of the 61 ABC genes in six common carp tissues (brain, heart, spleen, kidney, intestine, and gill) revealed extensive functional divergence among the ABC genes. Different copies of some genes had tissue-specific expression patterns, which may indicate some gene function specialization. This study provides essential genomic resources for future studies in common carp.

  18. Human ATP-binding cassette transporter 1 (ABC1): Genomic organization and identification of the genetic defect in the original Tangier disease kindred

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    Remaley, Alan T.; Rust, Stephan; Rosier, Marie; Knapper, Cathy; Naudin, Laurent; Broccardo, Cyril; Peterson, Katherine M.; Koch, Christine; Arnould, Isabelle; Prades, Catherine; Duverger, Nicholas; Funke, Harald; Assman, Gerd; Dinger, Maria; Dean, Michael; Chimini, Giovanna; Santamarina-Fojo, Silvia; Fredrickson, Donald S.; Denefle, Patrice; Brewer, H. Bryan

    1999-01-01

    Tangier disease is characterized by low serum high density lipoproteins and a biochemical defect in the cellular efflux of lipids to high density lipoproteins. ABC1, a member of the ATP-binding cassette family, recently has been identified as the defective gene in Tangier disease. We report here the organization of the human ABC1 gene and the identification of a mutation in the ABC1 gene from the original Tangier disease kindred. The organization of the human ABC1 gene is similar to that of the mouse ABC1 gene and other related ABC genes. The ABC1 gene contains 49 exons that range in size from 33 to 249 bp and is over 70 kb in length. Sequence analysis of the ABC1 gene revealed that the proband for Tangier disease was homozygous for a deletion of nucleotides 3283 and 3284 (TC) in exon 22. The deletion results in a frameshift mutation and a premature stop codon starting at nucleotide 3375. The product is predicted to encode a nonfunctional protein of 1,084 aa, which is approximately half the size of the full-length ABC1 protein. The loss of a Mnl1 restriction site, which results from the deletion, was used to establish the genotype of the rest of the kindred. In summary, we report on the genomic organization of the human ABC1 gene and identify a frameshift mutation in the ABC1 gene of the index case of Tangier disease. These results will be useful in the future characterization of the structure and function of the ABC1 gene and the analysis of additional ABC1 mutations in patients with Tangier disease. PMID:10535983

  19. Whole-Transcriptome Survey of the Putative ATP-Binding Cassette (ABC) Transporter Family Genes in the Latex-Producing Laticifers of Hevea brasiliensis

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    Zhiyi, Nie; Guijuan, Kang; Yu, Li; Longjun, Dai; Rizhong, Zeng

    2015-01-01

    The ATP-binding cassette (ABC) proteins or transporters constitute a large protein family in plants and are involved in many different cellular functions and processes, including solute transportation, channel regulation and molecular switches, etc. Through transcriptome sequencing, a transcriptome-wide survey and expression analysis of the ABC protein genes were carried out using the laticiferous latex from Hevea brasiliensis (rubber tree). A total of 46 putative ABC family proteins were identified in the H. brasiliensis latex. These consisted of 12 ‘full-size’, 21 ‘half-size’ and 13 other putative ABC proteins, and all of them showed strong conservation with their Arabidopsis thaliana counterparts. This study indicated that all eight plant ABC protein paralog subfamilies were identified in the H. brasiliensis latex, of which ABCB, ABCG and ABCI were the most abundant. Real-time quantitative reverse transcription-polymerase chain reaction assays demonstrated that gene expression of several latex ABC proteins was regulated by ethylene, jasmonic acid or bark tapping (a wound stress) stimulation, and that HbABCB15, HbABCB19, HbABCD1 and HbABCG21 responded most significantly of all to the abiotic stresses. The identification and expression analysis of the latex ABC family proteins could facilitate further investigation into their physiological involvement in latex metabolism and rubber biosynthesis by H. brasiliensis. PMID:25615936

  20. The function of the ATP-binding cassette (ABC) transporter ABCB1 is not susceptible to actin disruption

    NARCIS (Netherlands)

    Meszaros, Peter; Hummel, Ina; Klappe, Karin; Draghiciu, Oana; Hoekstra, Dick; Kok, Jan W.

    Previously we have shown that the activity of the multidrug transporter ABCC1 (multidrug resistance protein 1), and its localization in lipid rafts, depends on cortical actin (Hummel I, Klappe K, Ercan C, Kok JW. Mol. Pharm. 2011 79, 229-40). Here we show that the efflux activity of the ATP-binding

  1. Regulatory pathways for ATP-binding cassette transport proteins in kidney proximal tubules

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    Masereeuw, R.; Russel, F.G.M.

    2012-01-01

    The ATP-binding cassette transport proteins (ABC transporters) represent important determinants of drug excretion. Protective or excretory tissues where these transporters mediate substrate efflux include the kidney proximal tubule. Regulation of the transport proteins in this tissue requires

  2. Atovaquone and quinine anti-malarials inhibit ATP binding cassette transporter activity

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    Rijpma, S.R.; Heuvel, J.J.; Velden, M. van der; Sauerwein, R.W.; Russel, F.G.; Koenderink, J.B.

    2014-01-01

    BACKGROUND: Therapeutic blood plasma concentrations of anti-malarial drugs are essential for successful treatment. Pharmacokinetics of pharmaceutical compounds are dependent of adsorption, distribution, metabolism, and excretion. ATP binding cassette (ABC) transport proteins are particularly

  3. Genome-wide analysis of ATP-binding cassette (ABC) transporters in the sweetpotato whitefly, Bemisia tabaci.

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    Tian, Lixia; Song, Tianxue; He, Rongjun; Zeng, Yang; Xie, Wen; Wu, Qingjun; Wang, Shaoli; Zhou, Xuguo; Zhang, Youjun

    2017-04-26

    ABC transporter superfamily is one of the largest and ubiquitous groups of proteins. Because of their role in detoxification, insect ABC transporters have gained more attention in recent years. In this study, we annotated ABC transporters from a newly sequenced sweetpotato whitefly genome. Bemisia tabaci Q biotype is an emerging global invasive species that has caused extensive damages to field crops as well as ornamental plants. A total of 55 ABC transporters containing all eight described subfamilies (A to H) were identified in the B. tabaci Q genome, including 8 ABCAs, 3 ABCBs, 6 ABCCs, 2 ABCDs, 1 ABCE, 3 ABCFs, 23 ABCGs and 9 ABCHs. In comparison to other species, subfamilies G and H in both phloem- and blood-sucking arthropods are expanded. The temporal expression profiles of these 55 ABC transporters throughout B. tabaci developmental stages and their responses to imidacloprid, a neonicotinoid insecticide, were investigated using RNA-seq analysis. Furthermore, the mRNA expression of 24 ABC transporters (44% of the total) representing all eight subfamilies was confirmed by the quantitative real-time PCR (RT-qPCR). Furthermore, mRNA expression levels estimated by RT-qPCR and RNA-seq analyses were significantly correlated (r = 0.684, p ABC transporters in B. tabaci. The identification of these ABC transporters, their temporal expression profiles during B. tabaci development, and their response to a neonicotinoid insecticide lay the foundation for functional genomic understanding of their contribution to the invasiveness of B. tabaci.

  4. A mutation within the extended X loop abolished substrate-induced ATPase activity of the human liver ATP-binding cassette (ABC) transporter MDR3.

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    Kluth, Marianne; Stindt, Jan; Dröge, Carola; Linnemann, Doris; Kubitz, Ralf; Schmitt, Lutz

    2015-02-20

    The human multidrug resistance protein 3 (MDR3/ABCB4) belongs to the ubiquitous family of ATP-binding cassette (ABC) transporters and is located in the canalicular membrane of hepatocytes. There it flops the phospholipids of the phosphatidylcholine (PC) family from the inner to the outer leaflet. Here, we report the characterization of wild type MDR3 and the Q1174E mutant, which was identified previously in a patient with progressive familial intrahepatic cholestasis type 3 (PFIC-3). We expressed different variants of MDR3 in the yeast Pichia pastoris, purified the proteins via tandem affinity chromatography, and determined MDR3-specific ATPase activity in the presence or absence of phospholipids. The ATPase activity of wild type MDR3 was stimulated 2-fold by liver PC or 1,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine lipids. Furthermore, the cross-linking of MDR3 with a thiol-reactive fluorophore blocked ATP hydrolysis and exhibited no PC stimulation. Similarly, phosphatidylethanolamine, phosphatidylserine, and sphingomyelin lipids did not induce an increase of wild type MDR3 ATPase activity. The phosphate analogues beryllium fluoride and aluminum fluoride led to complete inhibition of ATPase activity, whereas orthovanadate inhibited exclusively the PC-stimulated ATPase activity of MDR3. The Q1174E mutation is located in the nucleotide-binding domain in direct proximity of the leucine of the ABC signature motif and extended the X loop, which is found in ABC exporters. Our data on the Q1174E mutant demonstrated basal ATPase activity, but PC lipids were incapable of stimulating ATPase activity highlighting the role of the extended X loop in the cross-talk of the nucleotide-binding domain and the transmembrane domain. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  5. Characterization of a lactose-responsive promoter of ATP-binding cassette (ABC) transporter gene from Lactobacillus acidophilus 05-172.

    Science.gov (United States)

    Zeng, Zhu; Zuo, Fanglei; Yu, Rui; Zhang, Bo; Ma, Huiqin; Chen, Shangwu

    2017-09-01

    A novel lactose-responsive promoter of the ATP-binding cassette (ABC) transporter gene Lba1680 of Lactobacillus acidophilus strain 05-172 isolated from a traditionally fermented dairy product koumiss was characterized. In L. acidophilus 05-172, expression of Lba1680 was induced by lactose, with lactose-induced transcription of Lba1680 being 6.1-fold higher than that induced by glucose. This is in contrast to L. acidophilus NCFM, a strain isolated from human feces, in which expression of Lba1680 and Lba1679 is induced by glucose. Both gene expression and enzyme activity assays in L. paracasei transformed with a vector containing the inducible Lba1680 promoter (PLba1680) of strain 05-172 and a heme-dependent catalase gene as reporter confirmed that PLba1680 is specifically induced by lactose. Its regulatory expression could not be repressed by glucose, and was independent of cAMP receptor protein. This lactose-responsive promoter might be used in the expression of functional genes in L. paracasei incorporated into a lactose-rich environment, such as dairy products. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  6. RCN1/OsABCG5, an ATP-binding cassette (ABC) transporter, is required for hypodermal suberization of roots in rice (Oryza sativa).

    Science.gov (United States)

    Shiono, Katsuhiro; Ando, Miho; Nishiuchi, Shunsaku; Takahashi, Hirokazu; Watanabe, Kohtaro; Nakamura, Motoaki; Matsuo, Yuichi; Yasuno, Naoko; Yamanouchi, Utako; Fujimoto, Masaru; Takanashi, Hideki; Ranathunge, Kosala; Franke, Rochus B; Shitan, Nobukazu; Nishizawa, Naoko K; Takamure, Itsuro; Yano, Masahiro; Tsutsumi, Nobuhiro; Schreiber, Lukas; Yazaki, Kazufumi; Nakazono, Mikio; Kato, Kiyoaki

    2014-10-01

    Suberin is a complex polymer composed of aliphatic and phenolic compounds. It is a constituent of apoplastic plant interfaces. In many plant species, including rice (Oryza sativa), the hypodermis in the outer part of roots forms a suberized cell wall (the Casparian strip and/or suberin lamellae), which inhibits the flow of water and ions and protects against pathogens. To date, there is no genetic evidence that suberin forms an apoplastic transport barrier in the hypodermis. We discovered that a rice reduced culm number1 (rcn1) mutant could not develop roots longer than 100 mm in waterlogged soil. The mutated gene encoded an ATP-binding cassette (ABC) transporter named RCN1/OsABCG5. RCN1/OsABCG5 gene expression in the wild type was increased in most hypodermal and some endodermal roots cells under stagnant deoxygenated conditions. A GFP-RCN1/OsABCG5 fusion protein localized at the plasma membrane of the wild type. Under stagnant deoxygenated conditions, well suberized hypodermis developed in wild types but not in rcn1 mutants. Under stagnant deoxygenated conditions, apoplastic tracers (periodic acid and berberine) were blocked at the hypodermis in the wild type but not in rcn1, indicating that the apoplastic barrier in the mutant was impaired. The amount of the major aliphatic suberin monomers originating from C(28) and C(30) fatty acids or ω-OH fatty acids was much lower in rcn1 than in the wild type. These findings suggest that RCN1/OsABCG5 has a role in the suberization of the hypodermis of rice roots, which contributes to formation of the apoplastic barrier. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

  7. Serum albumin promotes ATP-binding cassette transporter-dependent sterol uptake in yeast

    DEFF Research Database (Denmark)

    Marek, Magdalena; Silvestro, Daniele; Fredslund, Maria D.

    2014-01-01

    Sterol uptake in fungi is a multistep process that involves interaction between external sterols and the cell wall, incorporation of sterol molecules into the plasma membrane, and subsequent integration into intracellular membranes for turnover. ATP-binding cassette (ABC) transporters have been i...

  8. Metabolism of ATP-binding cassette drug transporter inhibitors: complicating factor for multidrug resistance.

    NARCIS (Netherlands)

    Cnubben, N.H.; Wortelboer, H.M.; Zanden, J.J. van; Rietjens, I.M.; Bladeren, P.J. van

    2005-01-01

    Membrane transport proteins belonging to the ATP-binding cassette (ABC) family of transport proteins play a central role in the defence of organisms against toxic compounds, including anticancer drugs. However, for compounds that are designed to display a toxic effect, this defence system diminishes

  9. ATP-binding cassette transporters in reproduction: a new frontier

    Science.gov (United States)

    Bloise, E.; Ortiga-Carvalho, T.M.; Reis, F.M.; Lye, S.J.; Gibb, W.; Matthews, S.G.

    2016-01-01

    BACKGROUND The transmembrane ATP-binding cassette (ABC) transporters actively efflux an array of clinically relevant compounds across biological barriers, and modulate biodistribution of many physiological and pharmacological factors. To date, over 48 ABC transporters have been identified and shown to be directly and indirectly involved in peri-implantation events and fetal/placental development. They efflux cholesterol, steroid hormones, vitamins, cytokines, chemokines, prostaglandins, diverse xenobiotics and environmental toxins, playing a critical role in regulating drug disposition, immunological responses and lipid trafficking, as well as preventing fetal accumulation of drugs and environmental toxins. METHODS This review examines ABC transporters as important mediators of placental barrier functions and key reproductive processes. Expression, localization and function of all identified ABC transporters were systematically reviewed using PubMed and Google Scholar websites to identify relevant studies examining ABC transporters in reproductive tissues in physiological and pathophysiological states. Only reports written in English were incorporated with no restriction on year of publication. While a major focus has been placed on the human, extensive evidence from animal studies is utilized to describe current understanding of the regulation and function of ABC transporters relevant to human reproduction. RESULTS ABC transporters are modulators of steroidogenesis, fertilization, implantation, nutrient transport and immunological responses, and function as ‘gatekeepers’ at various barrier sites (i.e. blood-testes barrier and placenta) against potentially harmful xenobiotic factors, including drugs and environmental toxins. These roles appear to be species dependent and change as a function of gestation and development. The best-described ABC transporters in reproductive tissues (primarily in the placenta) are the multidrug transporters p-glycoprotein and

  10. ATP-Binding Cassette Systems of Brucella

    Directory of Open Access Journals (Sweden)

    Dominic C. Jenner

    2009-01-01

    Full Text Available Brucellosis is a prevalent zoonotic disease and is endemic in the Middle East, South America, and other areas of the world. In this study, complete inventories of putative functional ABC systems of five Brucella species have been compiled and compared. ABC systems of Brucella melitensis 16M, Brucella abortus 9-941, Brucella canis RM6/66, Brucella suis 1330, and Brucella ovis 63/290 were identified and aligned. High numbers of ABC systems, particularly nutrient importers, were found in all Brucella species. However, differences in the total numbers of ABC systems were identified (B. melitensis, 79; B. suis, 72; B. abortus 64; B. canis, 74; B. ovis, 59 as well as specific differences in the functional ABC systems of the Brucella species. Since B. ovis is not known to cause human brucellosis, functional ABC systems absent in the B. ovis genome may represent virulence factors in human brucellosis.

  11. Genome-Wide Identification and Expression Profiling of ATP-Binding Cassette (ABC Transporter Gene Family in Pineapple (Ananas comosus (L. Merr. Reveal the Role of AcABCG38 in Pollen Development

    Directory of Open Access Journals (Sweden)

    Piaojuan Chen

    2017-12-01

    Full Text Available Pineapple (Ananas comosus L. cultivation commonly relies on asexual reproduction which is easily impeded by many factors in agriculture production. Sexual reproduction might be a novel approach to improve the pineapple planting. However, genes controlling pineapple sexual reproduction are still remain elusive. In different organisms a conserved superfamily proteins known as ATP binding cassette (ABC participate in various biological processes. Whereas, till today the ABC gene family has not been identified in pineapple. Here 100 ABC genes were identified in the pineapple genome and grouped into eight subfamilies (5 ABCAs, 20 ABCBs, 16 ABCCs, 2 ABCDs, one ABCEs, 5 ABCFs, 42 ABCGs and 9 ABCIs. Gene expression profiling revealed the dynamic expression pattern of ABC gene family in various tissues and different developmental stages. AcABCA5, AcABCB6, AcABCC4, AcABCC7, AcABCC9, AcABCG26, AcABCG38 and AcABCG42 exhibited preferential expression in ovule and stamen. Over-expression of AcABCG38 in the Arabidopsis double mutant abcg1-2abcg16-2 partially restored its pollen abortion defects, indicating that AcABCG38 plays important roles in pollen development. Our study on ABC gene family in pineapple provides useful information for developing sexual pineapple plantation which could be utilized to improve pineapple agricultural production.

  12. The Yeast ATP-binding Cassette (ABC) Transporter Ycf1p Enhances the Recruitment of the Soluble SNARE Vam7p to Vacuoles for Efficient Membrane Fusion*

    Science.gov (United States)

    Sasser, Terry L.; Lawrence, Gus; Karunakaran, Surya; Brown, Christopher; Fratti, Rutilio A.

    2013-01-01

    The Saccharomyces cerevisiae vacuole contains five ATP-binding cassette class C (ABCC) transporters, including Ycf1p, a family member that was originally characterized as a Cd2+ transporter. Ycf1p has also been found to physically interact with a wide array of proteins, including factors that regulate vacuole homeostasis. In this study, we examined the role of Ycf1p and other ABCC transporters in the regulation of vacuole homotypic fusion. We found that deletion of YCF1 attenuated in vitro vacuole fusion by up to 40% relative to wild-type vacuoles. Plasmid-expressed wild-type Ycf1p rescued the deletion phenotype; however, Ycf1p containing a mutation of the conserved Lys-669 to Met in the Walker A box of the first nucleotide-binding domain (Ycf1pK669M) was unable to complement the fusion defect of ycf1Δ vacuoles. This indicates that the ATPase activity of Ycf1p is required for its function in regulating fusion. In addition, we found that deleting YCF1 caused a striking decrease in vacuolar levels of the soluble SNARE Vam7p, whereas total cellular levels were not altered. The attenuated fusion of ycf1Δ vacuoles was rescued by the addition of recombinant Vam7p to in vitro experiments. Thus, Ycf1p contributes in the recruitment of Vam7p to the vacuole for efficient membrane fusion. PMID:23658021

  13. ATP-binding Cassette (ABC) Transport System Solute-binding Protein-guided Identification of Novel d-Altritol and Galactitol Catabolic Pathways in Agrobacterium tumefaciens C58.

    Science.gov (United States)

    Wichelecki, Daniel J; Vetting, Matthew W; Chou, Liyushang; Al-Obaidi, Nawar; Bouvier, Jason T; Almo, Steven C; Gerlt, John A

    2015-11-27

    Innovations in the discovery of the functions of uncharacterized proteins/enzymes have become increasingly important as advances in sequencing technology flood protein databases with an exponentially growing number of open reading frames. This study documents one such innovation developed by the Enzyme Function Initiative (EFI; U54GM093342), the use of solute-binding proteins for transport systems to identify novel metabolic pathways. In a previous study, this strategy was applied to the tripartite ATP-independent periplasmic transporters. Here, we apply this strategy to the ATP-binding cassette transporters and report the discovery of novel catabolic pathways for d-altritol and galactitol in Agrobacterium tumefaciens C58. These efforts resulted in the description of three novel enzymatic reactions as follows: 1) oxidation of d-altritol to d-tagatose via a dehydrogenase in Pfam family PF00107, a previously unknown reaction; 2) phosphorylation of d-tagatose to d-tagatose 6-phosphate via a kinase in Pfam family PF00294, a previously orphan EC number; and 3) epimerization of d-tagatose 6-phosphate C-4 to d-fructose 6-phosphate via a member of Pfam family PF08013, another previously unknown reaction. The epimerization reaction catalyzed by a member of PF08013 is especially noteworthy, because the functions of members of PF08013 have been unknown. These discoveries were assisted by the following two synergistic bioinformatics web tools made available by the Enzyme Function Initiative: the EFI-Enzyme Similarity Tool and the EFI-Genome Neighborhood Tool. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  14. ATP-binding Cassette (ABC) Transport System Solute-binding Protein-guided Identification of Novel d-Altritol and Galactitol Catabolic Pathways in Agrobacterium tumefaciens C58*

    Science.gov (United States)

    Wichelecki, Daniel J.; Vetting, Matthew W.; Chou, Liyushang; Al-Obaidi, Nawar; Bouvier, Jason T.; Almo, Steven C.; Gerlt, John A.

    2015-01-01

    Innovations in the discovery of the functions of uncharacterized proteins/enzymes have become increasingly important as advances in sequencing technology flood protein databases with an exponentially growing number of open reading frames. This study documents one such innovation developed by the Enzyme Function Initiative (EFI; U54GM093342), the use of solute-binding proteins for transport systems to identify novel metabolic pathways. In a previous study, this strategy was applied to the tripartite ATP-independent periplasmic transporters. Here, we apply this strategy to the ATP-binding cassette transporters and report the discovery of novel catabolic pathways for d-altritol and galactitol in Agrobacterium tumefaciens C58. These efforts resulted in the description of three novel enzymatic reactions as follows: 1) oxidation of d-altritol to d-tagatose via a dehydrogenase in Pfam family PF00107, a previously unknown reaction; 2) phosphorylation of d-tagatose to d-tagatose 6-phosphate via a kinase in Pfam family PF00294, a previously orphan EC number; and 3) epimerization of d-tagatose 6-phosphate C-4 to d-fructose 6-phosphate via a member of Pfam family PF08013, another previously unknown reaction. The epimerization reaction catalyzed by a member of PF08013 is especially noteworthy, because the functions of members of PF08013 have been unknown. These discoveries were assisted by the following two synergistic bioinformatics web tools made available by the Enzyme Function Initiative: the EFI-Enzyme Similarity Tool and the EFI-Genome Neighborhood Tool. PMID:26472925

  15. Molecular cloning and characterisation of three new ATP-binding cassette transporter genes from the wheat pathogen Mycosphaerella graminicola

    NARCIS (Netherlands)

    Stergiopoulos, I.; Gielkens, M.M.C.; Goodall, S.D.; Venema, K.; Waard, De M.A.

    2002-01-01

    Three single copy ATP-binding cassette (ABC) transporter encoding genes, designated MgAtr3, MgAtr4, and MgAtr5, were cloned and sequenced from the plant pathogenic fungus Mycosphaerella graminicola. The encoded ABC proteins all exhibit the [NBD-TMS6]2 configuration and can be classified as novel

  16. ROLE OF ATP BINDING CASSETTE SUB-FAMILY MEMBER 2 (ABCG2) IN MOUSE EMBRYONIC STEM CELL DEVELOPMENT.

    Science.gov (United States)

    ATP binding cassette sub-family member 2 (ABCG2), is a member of the ABC transporter superfamily and a principal xenobiotic transporter. ABCG2 is also highly expressed in certain stem cell populations where it is thought to be related to stem cell plasticity, although the role o...

  17. ATP-Binding Cassette Proteins: Towards a Computational View of Mechanism

    Science.gov (United States)

    Liao, Jielou

    2004-03-01

    Many large machine proteins can generate mechanical force and undergo large-scale conformational changes (LSCC) to perform varying biological tasks in living cells by utilizing ATP. Important examples include ATP-binding cassette (ABC) transporters. They are membrane proteins that couple ATP binding and hydrolysis to the translocation of substrates across membranes [1]. To interpret how the mechanical force generated by ATP binding and hydrolysis is propagated, a coarse-grained ATP-dependent harmonic network model (HNM) [2,3] is applied to the ABC protein, BtuCD. This protein machine transports vitamin B12 across membranes. The analysis shows that subunits of the protein move against each other in a concerted manner. The lowest-frequency modes of the BtuCD protein are found to link the functionally critical domains, and are suggested to be responsible for large-scale ATP-coupled conformational changes. [1] K. P. Locher, A. T. Lee and D. C. Rees. Science 296, 1091-1098 (2002). [2] Atilgan, A. R., S. R. Durell, R. L. Jernigan, M. C. Demirel, O. Keskin, and I. Bahar. Biophys. J. 80, 505-515(2002); M. M Tirion, Phys. Rev. Lett. 77, 1905-1908 (1996). [3] J. -L. Liao and D. N. Beratan, 2003, to be published.

  18. Membrane porters of ATP-binding cassette transport systems are polyphyletic.

    Science.gov (United States)

    Wang, Bin; Dukarevich, Maxim; Sun, Eric I; Yen, Ming Ren; Saier, Milton H

    2009-09-01

    The ATP-binding cassette (ABC) superfamily consists of both importers and exporters. These transporters have, by tradition, been classified according to the ATP hydrolyzing constituents, which are monophyletic. The evolutionary origins of the transmembrane porter proteins/domains are not known. Using five distinct computer programs, we here provide convincing statistical data suggesting that the transmembrane domains of ABC exporters are polyphyletic, having arisen at least three times independently. ABC1 porters arose by intragenic triplication of a primordial two-transmembrane segment (TMS)-encoding genetic element, yielding six TMS proteins. ABC2 porters arose by intragenic duplication of a dissimilar primordial three-TMS-encoding genetic element, yielding a distinctive protein family, nonhomologous to the ABC1 proteins. ABC3 porters arose by duplication of a primordial four-TMS-encoding genetic element, yielding either eight- or 10-TMS proteins. We assign each of 48 of the 50 currently recognized families of ABC exporters to one of the three evolutionarily distinct ABC types. Currently available high-resolution structural data for ABC porters are fully consistent with our findings. These results provide guides for future structural and mechanistic studies of these important transport systems.

  19. Cell and molecular biology of ATP-binding cassette proteins in plants.

    Science.gov (United States)

    Yazaki, Kazufumi; Shitan, Nobukazu; Sugiyama, Akifumi; Takanashi, Kojiro

    2009-01-01

    ATP-binding cassette (ABC) proteins constitute a large and diverse superfamily of membrane-bound and soluble proteins, which are involved in a wide range of biological processes in all organisms from prokaryotes to eukaryotes. Genome analyses of model plants, for example, Arabidopsis and rice, have revealed that plants have more than double numbers of this family member in their genomes compared to animals and insects. In recent years, various biochemical and physiological functions of ABC proteins in plants have been reported. Some are relevant for the defense mechanisms to biotic and abiotic stresses, whereas others are involved in the basic functions necessary for maintaining the plant life. Here, we provide an updated inventory of plant ABC proteins and summarize their tissue specificities, membrane localizations, and physiological functions.

  20. Association of ATP-binding cassette transporter-A1 polymorphism ...

    Indian Academy of Sciences (India)

    [Halalkhor S., Mesbah-Namin S. A., Daneshpour M. S., Hedayati M. and Azizi F. 2011 Association of ATP-binding cassette transporter-A1 polymorphism with apolipoprotein AI level in Tehranian population. J. Genet. 90, 129–132 ]. Introduction. The ATP-binding cassette transporter-A1 (ABCA1) plays a crucial role in reverse ...

  1. ATP-binding cassette systems in Burkholderia pseudomallei and Burkholderia mallei

    Directory of Open Access Journals (Sweden)

    Titball Richard W

    2007-03-01

    Full Text Available Abstract Background ATP binding cassette (ABC systems are responsible for the import and export of a wide variety of molecules across cell membranes and comprise one of largest protein superfamilies found in prokarya, eukarya and archea. ABC systems play important roles in bacterial lifestyle, virulence and survival. In this study, an inventory of the ABC systems of Burkholderia pseudomallei strain K96243 and Burkholderia mallei strain ATCC 23344 has been compiled using bioinformatic techniques. Results The ABC systems in the genomes of B. pseudomallei and B. mallei have been reannotated and subsequently compared. Differences in the number and types of encoded ABC systems in belonging to these organisms have been identified. For example, ABC systems involved in iron acquisition appear to be correlated with differences in genome size and lifestyles between these two closely related organisms. Conclusion The availability of complete inventories of the ABC systems in B. pseudomallei and B. mallei has enabled a more detailed comparison of the encoded proteins in this family. This has resulted in the identification of ABC systems which may play key roles in the different lifestyles and pathogenic properties of these two bacteria. This information has the potential to be exploited for improved clinical identification of these organisms as well as in the development of new vaccines and therapeutics targeted against the diseases caused by these organisms.

  2. Regulatory pathways for ATP-binding cassette transport proteins in kidney proximal tubules.

    Science.gov (United States)

    Masereeuw, Rosalinde; Russel, Frans G M

    2012-12-01

    The ATP-binding cassette transport proteins (ABC transporters) represent important determinants of drug excretion. Protective or excretory tissues where these transporters mediate substrate efflux include the kidney proximal tubule. Regulation of the transport proteins in this tissue requires elaborate signaling pathways, including genetic, epigenetic, nuclear receptor mediated, posttranscriptional gene regulation involving microRNAs, and non-genomic (kinases) pathways triggered by hormones and/or growth factors. This review discusses current knowledge on regulatory pathways for ABC transporters in kidney proximal tubules, with a main focus on P-glycoprotein, multidrug resistance proteins 2 and 4, and breast cancer resistance protein. Insight in these processes is of importance because variations in transporter activity due to certain (disease) conditions could lead to significant changes in drug efficacy or toxicity.

  3. LrABCF1, a GCN-type ATP-binding cassette transporter from lilium regale, is involved in defense responses against viral and fungal pathogens

    Science.gov (United States)

    ATP-binding cassette (ABC) transporters are essential for membrane translocation in diverse biological processes, such as plant development and defense response. Here, a general control non-derepressible (GCN)-type ABC transporter gene, designated LrABCF1, was identified from Cucumber mosaic virus (...

  4. Expression of some ATP-binding cassette transporters in acute myeloid leukemia

    Directory of Open Access Journals (Sweden)

    Antonella Maria Salvia

    2017-12-01

    Full Text Available Hematopoietic cells express ATP binding cassette (ABC transporters in relation to different degrees of differentiation. One of the known multidrug resistance mechanisms in acute myeloid leukemia (AML is the overexpression of efflux pumps belonging to the superfamily of ABC transporters such as ABCB1, ABCG2 and ABCC1. Although several studies were carried out to correlate ABC transporters expression with drug resistance, little is known about their role as markers of diagnosis and progression of the disease. For this purpose we investigated the expression, by real-time PCR, of some ABC genes in bone marrow samples of AML patients at diagnosis and after induction therapy. At diagnosis, ABCG2 was always down-regulated, while an up regulated trend for ABCC1 was observed. After therapy the examined genes showed a different expression trend and approached the values of healthy subjects suggesting that this event could be considered as a marker of AML regression. The expression levels of some ABC transporters such as ABCC6, seems to be related to gender, age and to the presence of FLT3/ITD gene mutation.

  5. ATP-binding cassette transporters ABCA1, ABCA7, and ABCG1 in mouse spermatozoa.

    Science.gov (United States)

    Morales, Carlos R; Marat, Andrea L; Ni, Xiaoyan; Yu, Yang; Oko, Richard; Smith, Brian T; Argraves, W Scott

    2008-11-21

    Mammalian spermatozoa lose plasma membrane cholesterol during their maturation in the epididymis and during their capacitation in the female reproductive tract. While acceptors such as high-density lipoproteins (HDL) and apolipoproteins A-I (apoA-I) and J have been found in male and female reproductive tracts, transporters that mediate cholesterol efflux from plasma membranes of spermatozoa to such acceptors have not yet been defined. Candidate transporters are members of the ATP-binding cassette (ABC) transporter superfamily including ABCA1, ABCA7, ABCG1 and ABCG4, which have all been implicated in the transport of sterols and phospholipids to apolipoproteins and HDL. Here we show that mouse spermatozoa in the seminiferous tubules and epididymis express ABCA1, ABCA7 and ABCG1, but not ABCG4. Moreover, we show that ABCA1, ABCA7, and ABCG1 antibodies decrease cholesterol efflux from spermatozoa to lipid acceptors apoA-I and albumin and inhibit in vitro fertilization.

  6. ATP-binding cassette transporters of the multicellular cyanobacterium Anabaena sp. PCC 7120: a wide variety for a complex lifestyle.

    Science.gov (United States)

    Shvarev, Dmitry; Maldener, Iris

    2018-02-01

    Two hundred genes or 3% of the known or putative protein-coding genes of the filamentous freshwater cyanobacterium Anabaena sp. PCC 7120 encode domains of ATP-binding cassette (ABC) transporters. Detailed characterization of some of these transporters (14-15 importers and 5 exporters) has revealed their crucial roles in the complex lifestyle of this multicellular photoautotroph, which is able to differentiate specialized cells for nitrogen fixation. This review summarizes the characteristics of the ABC transporters of Anabaena sp. PCC 7120 known to date. © FEMS 2018. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  7. Relation between hepatic expression of ATP-binding cassette transporters G5 and G8 and biliary cholesterol secretion in mice

    NARCIS (Netherlands)

    Kosters, A; Frijters, RJJM; Schaap, FG; Vink, E; Plosch, T; Ottenhoff, R; Jirsa, M; De Cuyper, IM; Kuipers, F; Groen, AK

    Background/Aims: Mutations in genes encoding the ATP-binding cassette (ABC)-transporters ABCG5 and ABCG8 underlie sitosterolemia, which is characterized by elevated plasma levels of phytosterols due to increased intestinal absorption and impaired biliary secretion of sterols. The aim of our study

  8. Relation between hepatic expression of ATP-binding cassette transporters G5 and G8 and biliary cholesterol secretion in mice

    NARCIS (Netherlands)

    Kosters, Astrid; Frijters, Raoul J. J. M.; Schaap, Frank G.; Vink, Edwin; Plösch, Torsten; Ottenhoff, Roelof; Jirsa, Milan; de Cuyper, Iris M.; Kuipers, Folkert; Groen, Albert K.

    2003-01-01

    Background/Aims: Mutations in genes encoding the ATP-binding cassette (ABC)-transporters ABCG5 and ABCG8 underlie sitosterolemia, which is characterized by elevated plasma levels of phytosterols due to increased intestinal absorption and impaired biliary secretion of sterols. The aim of our study

  9. Maltose-binding protein effectively stabilizes the partially closed conformation of the ATP-binding cassette transporter MalFGK2

    KAUST Repository

    Weng, Jingwei

    2017-02-23

    Maltose transporter MalFGK2 is a type-I importer in the ATP-binding cassette (ABC) transporter superfamily. Upon the binding of its periplasmic binding protein, MalE, the ATPase activity of MalFGK2 can be greatly enhanced. Crystal structures of the MalFGK2-MalE-maltose complex in a so-called

  10. Endocrine Disruptors Differentially Target ATP-Binding Cassette Transporters in the Blood-Testis Barrier and Affect Leydig Cell Testosterone Secretion In Vitro

    NARCIS (Netherlands)

    Dankers, A.C.A.; Roelofs, M.J.; Piersma, A.H.; Sweep, F.C.; Russel, F.G.M.; Berg, M. van den; Duursen, M.B. van; Masereeuw, R.

    2013-01-01

    Endocrine-disrupting chemicals (EDCs) are considered to cause testicular toxicity primarily via interference with steroid hormone function. Alternatively, EDCs could possibly exert their effects by interaction with ATP-binding cassette (ABC) transporters that are expressed in the blood-testis

  11. ATP Binding Cassette Transporter Mediates Both Heme and Pesticide Detoxification in Tick Midgut Cells.

    Directory of Open Access Journals (Sweden)

    Flavio Alves Lara

    Full Text Available In ticks, the digestion of blood occurs intracellularly and proteolytic digestion of hemoglobin takes place in a dedicated type of lysosome, the digest vesicle, followed by transfer of the heme moiety of hemoglobin to a specialized organelle that accumulates large heme aggregates, called hemosomes. In the present work, we studied the uptake of fluorescent metalloporphyrins, used as heme analogs, and amitraz, one of the most regularly used acaricides to control cattle tick infestations, by Rhipicephalus (Boophilus microplus midgut cells. Both compounds were taken up by midgut cells in vitro and accumulated inside the hemosomes. Transport of both molecules was sensitive to cyclosporine A (CsA, a well-known inhibitor of ATP binding cassette (ABC transporters. Rhodamine 123, a fluorescent probe that is also a recognized ABC substrate, was similarly directed to the hemosome in a CsA-sensitive manner. Using an antibody against conserved domain of PgP-1-type ABC transporter, we were able to immunolocalize PgP-1 in the digest vesicle membranes. Comparison between two R. microplus strains that were resistant and susceptible to amitraz revealed that the resistant strain detoxified both amitraz and Sn-Pp IX more efficiently than the susceptible strain, a process that was also sensitive to CsA. A transcript containing an ABC transporter signature exhibited 2.5-fold increased expression in the amitraz-resistant strain when compared with the susceptible strain. RNAi-induced down-regulation of this ABC transporter led to the accumulation of metalloporphyrin in the digestive vacuole, interrupting heme traffic to the hemosome. This evidence further confirms that this transcript codes for a heme transporter. This is the first report of heme transport in a blood-feeding organism. While the primary physiological function of the hemosome is to detoxify heme and attenuate its toxicity, we suggest that the use of this acaricide detoxification pathway by ticks may

  12. ATP Binding Cassette Transporter Mediates Both Heme and Pesticide Detoxification in Tick Midgut Cells

    Science.gov (United States)

    Lara, Flavio Alves; Pohl, Paula C.; Gandara, Ana Caroline; Ferreira, Jessica da Silva; Nascimento-Silva, Maria Clara; Bechara, Gervásio Henrique; Sorgine, Marcos H. F.; Almeida, Igor C.; Vaz, Itabajara da Silva; Oliveira, Pedro L.

    2015-01-01

    In ticks, the digestion of blood occurs intracellularly and proteolytic digestion of hemoglobin takes place in a dedicated type of lysosome, the digest vesicle, followed by transfer of the heme moiety of hemoglobin to a specialized organelle that accumulates large heme aggregates, called hemosomes. In the present work, we studied the uptake of fluorescent metalloporphyrins, used as heme analogs, and amitraz, one of the most regularly used acaricides to control cattle tick infestations, by Rhipicephalus (Boophilus) microplus midgut cells. Both compounds were taken up by midgut cells in vitro and accumulated inside the hemosomes. Transport of both molecules was sensitive to cyclosporine A (CsA), a well-known inhibitor of ATP binding cassette (ABC) transporters. Rhodamine 123, a fluorescent probe that is also a recognized ABC substrate, was similarly directed to the hemosome in a CsA-sensitive manner. Using an antibody against conserved domain of PgP-1-type ABC transporter, we were able to immunolocalize PgP-1 in the digest vesicle membranes. Comparison between two R. microplus strains that were resistant and susceptible to amitraz revealed that the resistant strain detoxified both amitraz and Sn-Pp IX more efficiently than the susceptible strain, a process that was also sensitive to CsA. A transcript containing an ABC transporter signature exhibited 2.5-fold increased expression in the amitraz-resistant strain when compared with the susceptible strain. RNAi-induced down-regulation of this ABC transporter led to the accumulation of metalloporphyrin in the digestive vacuole, interrupting heme traffic to the hemosome. This evidence further confirms that this transcript codes for a heme transporter. This is the first report of heme transport in a blood-feeding organism. While the primary physiological function of the hemosome is to detoxify heme and attenuate its toxicity, we suggest that the use of this acaricide detoxification pathway by ticks may represent a new

  13. Whole-genome survey of the putative ATP-binding cassette transporter family genes in Vitis vinifera.

    Science.gov (United States)

    Çakır, Birsen; Kılıçkaya, Ozan

    2013-01-01

    The ATP-binding cassette (ABC) protein superfamily constitutes one of the largest protein families known in plants. In this report, we performed a complete inventory of ABC protein genes in Vitis vinifera, the whole genome of which has been sequenced. By comparison with ABC protein members of Arabidopsis thaliana, we identified 135 putative ABC proteins with 1 or 2 NBDs in V. vinifera. Of these, 120 encode intrinsic membrane proteins, and 15 encode proteins missing TMDs. V. vinifera ABC proteins can be divided into 13 subfamilies with 79 "full-size," 41 "half-size," and 15 "soluble" putative ABC proteins. The main feature of the Vitis ABC superfamily is the presence of 2 large subfamilies, ABCG (pleiotropic drug resistance and white-brown complex homolog) and ABCC (multidrug resistance-associated protein). We identified orthologs of V. vinifera putative ABC transporters in different species. This work represents the first complete inventory of ABC transporters in V. vinifera. The identification of Vitis ABC transporters and their comparative analysis with the Arabidopsis counterparts revealed a strong conservation between the 2 species. This inventory could help elucidate the biological and physiological functions of these transporters in V. vinifera.

  14. Whole-genome survey of the putative ATP-binding cassette transporter family genes in Vitis vinifera.

    Directory of Open Access Journals (Sweden)

    Birsen Çakır

    Full Text Available The ATP-binding cassette (ABC protein superfamily constitutes one of the largest protein families known in plants. In this report, we performed a complete inventory of ABC protein genes in Vitis vinifera, the whole genome of which has been sequenced. By comparison with ABC protein members of Arabidopsis thaliana, we identified 135 putative ABC proteins with 1 or 2 NBDs in V. vinifera. Of these, 120 encode intrinsic membrane proteins, and 15 encode proteins missing TMDs. V. vinifera ABC proteins can be divided into 13 subfamilies with 79 "full-size," 41 "half-size," and 15 "soluble" putative ABC proteins. The main feature of the Vitis ABC superfamily is the presence of 2 large subfamilies, ABCG (pleiotropic drug resistance and white-brown complex homolog and ABCC (multidrug resistance-associated protein. We identified orthologs of V. vinifera putative ABC transporters in different species. This work represents the first complete inventory of ABC transporters in V. vinifera. The identification of Vitis ABC transporters and their comparative analysis with the Arabidopsis counterparts revealed a strong conservation between the 2 species. This inventory could help elucidate the biological and physiological functions of these transporters in V. vinifera.

  15. A common highly conserved cadmium detoxification mechanism from bacteria to humans: heavy metal tolerance conferred by the ATP-binding cassette (ABC) transporter SpHMT1 requires glutathione but not metal-chelating phytochelatin peptides.

    Science.gov (United States)

    Prévéral, Sandra; Gayet, Landry; Moldes, Cristina; Hoffmann, Jonathan; Mounicou, Sandra; Gruet, Antoine; Reynaud, Florie; Lobinski, Ryszard; Verbavatz, Jean-Marc; Vavasseur, Alain; Forestier, Cyrille

    2009-02-20

    Cadmium poses a significant threat to human health due to its toxicity. In mammals and in bakers' yeast, cadmium is detoxified by ATP-binding cassette transporters after conjugation to glutathione. In fission yeast, phytochelatins constitute the co-substrate with cadmium for the transporter SpHMT1. In plants, a detoxification mechanism similar to the one in fission yeast is supposed, but the molecular nature of the transporter is still lacking. To investigate further the relationship between SpHMT1 and its co-substrate, we overexpressed the transporter in a Schizosaccharomyces pombe strain deleted for the phytochelatin synthase gene and heterologously in Saccharomyces cerevisiae and in Escherichia coli. In all organisms, overexpression of SpHMT1 conferred a markedly enhanced tolerance to cadmium but not to Sb(III), AgNO(3), As(III), As(V), CuSO(4), or HgCl(2). Abolishment of the catalytic activity by expression of SpHMT1(K623M) mutant suppressed the cadmium tolerance phenotype independently of the presence of phytochelatins. Depletion of the glutathione pool inhibited the SpHMT1 activity but not that of AtHMA4, a P-type ATPase, indicating that GSH is necessary for the SpHMT1-mediated cadmium resistance. In E. coli, SpHMT1 was targeted to the periplasmic membrane and led to an increased amount of cadmium in the periplasm. These results demonstrate that SpHMT1 confers cadmium tolerance in the absence of phytochelatins but depending on the presence of GSH and ATP. Our results challenge the dogma of the two separate cadmium detoxification pathways and demonstrate that a common highly conserved mechanism has been selected during the evolution from bacteria to humans.

  16. Structure-function analysis of peroxisomal ATP-binding cassette transporters using chimeric dimers

    NARCIS (Netherlands)

    Geillon, Flore; Gondcaille, Catherine; Charbonnier, Soëli; van Roermund, Carlo W.; Lopez, Tatiana E.; Dias, Alexandre M. M.; Pais de Barros, Jean-Paul; Arnould, Christine; Wanders, Ronald J.; Trompier, Doriane; Savary, Stéphane

    2014-01-01

    ABCD1 and ABCD2 are two closely related ATP-binding cassette half-transporters predicted to homodimerize and form peroxisomal importers for fatty acyl-CoAs. Available evidence has shown that ABCD1 and ABCD2 display a distinct but overlapping substrate specificity, although much remains to be learned

  17. ATP-binding cassette B10 regulates early steps of heme synthesis.

    Science.gov (United States)

    Bayeva, Marina; Khechaduri, Arineh; Wu, Rongxue; Burke, Michael A; Wasserstrom, J Andrew; Singh, Neha; Liesa, Marc; Shirihai, Orian S; Langer, Nathaniel B; Paw, Barry H; Ardehali, Hossein

    2013-07-19

    Heme plays a critical role in gas exchange, mitochondrial energy production, and antioxidant defense in cardiovascular system. The mitochondrial transporter ATP-binding cassette (ABC) B10 has been suggested to export heme out of the mitochondria and is required for normal hemoglobinization of erythropoietic cells and protection against ischemia-reperfusion injury in the heart; however, its primary function has not been established. The aim of this study was to identify the function of ABCB10 in heme synthesis in cardiac cells. Knockdown of ABCB10 in cardiac myoblasts significantly reduced heme levels and the activities of heme-containing proteins, whereas supplementation with δ-aminolevulinic acid reversed these defects. Overexpression of mitochondrial δ-aminolevulinic acid synthase 2, the rate-limiting enzyme upstream of δ-aminolevulinic acid export, failed to restore heme levels in cells with ABCB10 downregulation. ABCB10 and heme levels were increased by hypoxia, and reversal of ABCB10 upregulation caused oxidative stress and cell death. Furthermore, ABCB10 knockdown in neonatal rat cardiomyocytes resulted in a significant delay of calcium removal from the cytoplasm, suggesting a relaxation defect. Finally, ABCB10 expression and heme levels were altered in failing human hearts and mice with ischemic cardiomyopathy. ABCB10 plays a critical role in heme synthesis pathway by facilitating δ-aminolevulinic acid production or export from the mitochondria. In contrast to previous reports, we show that ABCB10 is not a heme exporter and instead is required for the early mitochondrial steps of heme biosynthesis.

  18. Structure, function, and evolution of bacterial ATP-binding cassette systems

    Energy Technology Data Exchange (ETDEWEB)

    Davidson, A.L.; Dassa, E.; Orelle, C.; Chen, J. (Purdue)

    2010-07-27

    The ATP-binding cassette (ABC) systems constitute one of the largest superfamilies of paralogous sequences. All ABC systems share a highly conserved ATP-hydrolyzing domain or protein (the ABC; also referred to as a nucleotide-binding domain [NBD]) that is unequivocally characterized by three short sequence motifs (Fig. 1): these are the Walker A and Walker B motifs, indicative of the presence of a nucleotide-binding site, and the signature motif, unique to ABC proteins, located upstream of the Walker B motif (426). Other motifs diagnostic of ABC proteins are also indicated in Fig. 1. The biological significance of these motifs is discussed in Structure, Function, and Dynamics of the ABC. ABC systems are widespread among living organisms and have been detected in all genera of the three kingdoms of life, with remarkable conservation in the primary sequence of the cassette and in the organization of the constitutive domains or subunits (203, 420). ABC systems couple the energy of ATP hydrolysis to an impressively large variety of essential biological phenomena, comprising not only transmembrane (TM) transport, for which they are best known, but also several non-transport-related processes, such as translation elongation (62) and DNA repair (174). Although ABC systems deserve much attention because they are involved in severe human inherited diseases (107), they were first discovered and characterized in detail in prokaryotes, as early as the 1970s (13, 148, 238, 468). The most extensively analyzed systems were the high-affinity histidine and maltose uptake systems of Salmonella enterica serovar Typhimurium and Escherichia coli. Over 2 decades ago, after the completion of the nucleotide sequences encoding these transporters in the respective laboratories of Giovanna Ames and Maurice Hofnung, Hiroshi Nikaido and colleagues noticed that the two systems displayed a global similarity in the nature of their components and, moreover, that the primary sequences of MalK and

  19. Localization of the ATP-binding cassette (ABC) transport proteins PfMRP1, PfMRP2, and PfMDR5 at the Plasmodium falciparum plasma membrane.

    NARCIS (Netherlands)

    Kavishe, R.A.; Heuvel, J.M.W. van den; Vegte-Bolmer, M.G. van de; Luty, A.J.F.; Russel, F.G.M.; Koenderink, J.B.

    2009-01-01

    BACKGROUND: The spread of drug resistance has been a major obstacle to the control of malaria. The mechanisms underlying drug resistance in malaria seem to be complex and multigenic. The current literature on multiple drug resistance against anti-malarials has documented PfMDR1, an ATP-binding

  20. Evaluation of the role of ATP-binding cassette transporters as a defence mechanism against temephos in populations of Aedes aegypti

    Directory of Open Access Journals (Sweden)

    Estelita Pereira Lima

    2014-11-01

    Full Text Available The role of ATP-binding cassette (ABC transporters in the efflux of the insecticide, temephos, was assessed in the larvae of Aedes aegypti. Bioassays were conducted using mosquito populations that were either susceptible or resistant to temephos by exposure to insecticide alone or in combination with sublethal doses of the ABC transporter inhibitor, verapamil (30, 35 and 40 μM. The best result in the series was obtained with the addition of verapamil (40 μM, which led to a 2x increase in the toxicity of temephos, suggesting that ABC transporters may be partially involved in conferring resistance to the populations evaluated.

  1. Atovaquone and quinine anti-malarials inhibit ATP binding cassette transporter activity.

    Science.gov (United States)

    Rijpma, Sanna R; van den Heuvel, Jeroen J M W; van der Velden, Maarten; Sauerwein, Robert W; Russel, Frans G M; Koenderink, Jan B

    2014-09-13

    Therapeutic blood plasma concentrations of anti-malarial drugs are essential for successful treatment. Pharmacokinetics of pharmaceutical compounds are dependent of adsorption, distribution, metabolism, and excretion. ATP binding cassette (ABC) transport proteins are particularly involved in drug deposition, as they are located at membranes of many uptake and excretory organs and at protective barriers, where they export endogenous and xenobiotic compounds, including pharmaceuticals. In this study, a panel of well-established anti-malarial drugs which may affect drug plasma concentrations was tested for interactions with human ABC transport proteins. The interaction of chloroquine, quinine, artemisinin, mefloquine, lumefantrine, atovaquone, dihydroartemisinin and proguanil, with transport activity of P-glycoprotein (P-gp), breast cancer resistance protein (BCRP), bile salt export pump (BSEP) and multidrug resistance-associated proteins (MRP) 1-4 were analysed. The effect of the anti-malarials on the ATP-dependent uptake of radio-labelled substrates was measured in membrane vesicles isolated from HEK293 cells overexpressing the ABC transport proteins. A strong and previously undescribed inhibition of BCRP-mediated transport by atovaquone with a 50% inhibitory concentration (IC50) of 0.23 μM (95% CI 0.17-0.29 μM) and inhibition of P-gp-mediated transport by quinine with an IC50 of 6.8 μM (95% CI 5.9-7.8 μM) was observed. Furthermore, chloroquine and mefloquine were found to significantly inhibit P-gp-mediated transport. BCRP transport activity was significantly inhibited by all anti-malarials tested, whereas BSEP-mediated transport was not inhibited by any of the compounds. Both MRP1- and MRP3-mediated transport were significantly inhibited by mefloquine. Atovaquone and quinine significantly inhibit BCRP- and P-gp- mediated transport at concentrations within the clinically relevant prophylactic and therapeutic range. Co-administration of these established anti

  2. Sorafenib modulates the gene expression of multi-drug resistance mediating ATP-binding cassette proteins in experimental hepatocellular carcinoma.

    Science.gov (United States)

    Hoffmann, Katrin; Franz, Clemens; Xiao, Zhi; Mohr, Elvira; Serba, Susanne; Büchler, Markus W; Schemmer, Peter

    2010-11-01

    High ATP-binding cassette (ABC) protein expression leads to intrinsic drug resistance of hepatocellular carcinoma (HCC). The aim of this study was to investigate the potential chemosensitizing effects of sorafenib on the multi-drug resistance (MDR) phenotype. The ABC-protein gene expression and the cellular survival were determined by RT-PCR analysis and MTT assay in HUH7 cells. Sorafenib inhibits MDR. The ABC-protein mRNA expression decreased by up to 51% (p ≤ 0.01). Addition of sorafenib to conventional chemotherapy restored the chemosensitivity. Combination of gemcitabine plus sorafenib decreased the ABC-protein mRNA levels by up to 77%, compared to gemcitabine monotherapy (p ≤ 0.001). Doxorubicin plus sorafenib decreased the ABC-protein mRNA levels up to 74% compared to doxorubicin monotherapy (p ≤ 0.001). This study provides evidence that the MDR phenotype of HCC cells can be modulated by the multi-kinase inhibitor sorafenib and consequentially may lead towards personalized therapies in patients with highly resistant tumors.

  3. Microarray study of single nucleotide polymorphisms and expression of ATP-binding cassette genes in breast tumors

    Science.gov (United States)

    Tsyganov, M. M.; Ibragimova, M. K.; Karabut, I. V.; Freydin, M. B.; Choinzonov, E. L.; Litvyakov, N. V.

    2015-11-01

    Our previous research establishes that changes of expression of the ATP-binding cassette genes family is connected with the neoadjuvant chemotherapy effect. However, the mechanism of regulation of resistance gene expression remains unclear. As many researchers believe, single nucleotide polymorphisms can be involved in this process. Thereupon, microarray analysis is used to study polymorphisms in ATP-binding cassette genes. It is thus found that MDR gene expression is connected with 5 polymorphisms, i.e. rs241432, rs241429, rs241430, rs3784867, rs59409230, which participate in the regulation of expression of own genes.

  4. Genetic Analysis of the Mode of Interplay between an ATPase Subunit and Membrane Subunits of the Lipoprotein-Releasing ATP-Binding Cassette Transporter LolCDE†

    OpenAIRE

    Ito, Yasuko; Matsuzawa, Hitomi; Matsuyama, Shin-ichi; Narita, Shin-ichiro; Tokuda, Hajime

    2006-01-01

    The LolCDE complex, an ATP-binding cassette (ABC) transporter, releases lipoproteins from the inner membrane, thereby initiating lipoprotein sorting to the outer membrane of Escherichia coli. The LolCDE complex is composed of two copies of an ATPase subunit, LolD, and one copy each of integral membrane subunits LolC and LolE. LolD hydrolyzes ATP on the cytoplasmic side of the inner membrane, while LolC and/or LolE recognize and release lipoproteins anchored to the periplasmic leaflet of the i...

  5. An ATP-binding cassette subfamily G full transporter is essential for the retention of leaf water in both wild barley and rice.

    OpenAIRE

    Chen Guoxiong; Komatsuda Takao; Ma Jian Feng; Nawrath Christiane; Pourkheirandish Mohammad; Tagiri Akemi; Hu Yin-Gang; Sameri Mohammad; Li Xinrong; Zhao Xin; Liu Yubing; Li Chao; Ma Xiaoying; Wang Aidong; Nair Sudha

    2011-01-01

    Land plants have developed a cuticle preventing uncontrolled water loss. Here we report that an ATP-binding cassette (ABC) subfamily G (ABCG) full transporter is required for leaf water conservation in both wild barley and rice. A spontaneous mutation, eibi1.b, in wild barley has a low capacity to retain leaf water, a phenotype associated with reduced cutin deposition and a thin cuticle. Map-based cloning revealed that Eibi1 encodes an HvABCG31 full transporter. The gene was highly expressed ...

  6. Functional analysis of an ATP-binding cassette transporter protein from Aspergillus fumigatus by heterologous expression in Saccharomyces cerevisiae.

    Science.gov (United States)

    Paul, Sanjoy; Moye-Rowley, W Scott

    2013-08-01

    Aspergillus fumigatus is the major filamentous fungal pathogen in humans. Although A. fumigatus can be treated with many of the available antifungal drugs, including azole compounds, drug resistant isolates are being recovered at an increasing rate. In other fungal pathogens such as the Candida species, ATP-binding cassette (ABC) transporter proteins play important roles in development of clinically-significant azole resistance phenotypes. Central among these ABC transporter proteins are homologues of the Saccharomyces cerevisiae Pdr5 multidrug transporter. In this work, we test the two A. fumigatus genes encoding proteins sharing the highest degree of sequence similarity to S. cerevisiae Pdr5 for their ability to be function in a heterologous pdr5Δ strain of S. cerevisiae. Expression of full-length cDNAs for these two Afu proteins failed to suppress the drug sensitive phenotype of a pdr5Δ strain and no evidence could be obtained for their expression as green fluorescent protein (GFP) fusions. To improve the expression of one of these Afu ABC transporters (XP_755847), we changed the sequence of the cDNA to use codons corresponding to the major tRNA species in S. cerevisiae. This codon-optimized (CO Afu abcA) cDNA was efficiently expressed in pdr5Δ cells and able to be detected as a GFP fusion protein. The CO Afu abcA did not correct the drug sensitivity of the pdr5Δ strain and exhibited a high degree of perinuclear fluorescence suggesting that this fusion protein was localized to the S. cerevisiae ER. Interestingly, when these experiments were repeated at 37 °C, the CO Afu abcA was able to complement the drug sensitive phenotype of pdr5Δ cells and exhibited less intracellular fluorescence. Additionally, we found that the CO Afu abcA was able to reduce resistance to drugs like phytosphingosine that act via causing mislocalization of amino acid permeases in fungi. These data suggest that the Afu abcA protein can carry out two different functions of Pdr5: drug

  7. Structure-Function Analysis of Peroxisomal ATP-binding Cassette Transporters Using Chimeric Dimers*

    Science.gov (United States)

    Geillon, Flore; Gondcaille, Catherine; Charbonnier, Soëli; Van Roermund, Carlo W.; Lopez, Tatiana E.; Dias, Alexandre M. M.; Pais de Barros, Jean-Paul; Arnould, Christine; Wanders, Ronald J.; Trompier, Doriane; Savary, Stéphane

    2014-01-01

    ABCD1 and ABCD2 are two closely related ATP-binding cassette half-transporters predicted to homodimerize and form peroxisomal importers for fatty acyl-CoAs. Available evidence has shown that ABCD1 and ABCD2 display a distinct but overlapping substrate specificity, although much remains to be learned in this respect as well as in their capability to form functional heterodimers. Using a cell model expressing an ABCD2-EGFP fusion protein, we first demonstrated by proximity ligation assay and co-immunoprecipitation assay that ABCD1 interacts with ABCD2. Next, we tested in the pxa1/pxa2Δ yeast mutant the functionality of ABCD1/ABCD2 dimers by expressing chimeric proteins mimicking homo- or heterodimers. For further structure-function analysis of ABCD1/ABCD2 dimers, we expressed chimeric dimers fused to enhanced GFP in human skin fibroblasts of X-linked adrenoleukodystrophy patients. These cells are devoid of ABCD1 and accumulate very long-chain fatty acids (C26:0 and C26:1). We checked that the chimeric proteins were correctly expressed and targeted to the peroxisomes. Very long-chain fatty acid levels were partially restored in transfected X-linked adrenoleukodystrophy fibroblasts regardless of the chimeric construct used, thus demonstrating functionality of both homo- and heterodimers. Interestingly, the level of C24:6 n-3, the immediate precursor of docosahexaenoic acid, was decreased in cells expressing chimeric proteins containing at least one ABCD2 moiety. Our data demonstrate for the first time that both homo- and heterodimers of ABCD1 and ABCD2 are functionally active. Interestingly, the role of ABCD2 (in homo- and heterodimeric forms) in the metabolism of polyunsaturated fatty acids is clearly evidenced, and the chimeric dimers provide a novel tool to study substrate specificity of peroxisomal ATP-binding cassette transporters. PMID:25043761

  8. Structure-function analysis of peroxisomal ATP-binding cassette transporters using chimeric dimers.

    Science.gov (United States)

    Geillon, Flore; Gondcaille, Catherine; Charbonnier, Soëli; Van Roermund, Carlo W; Lopez, Tatiana E; Dias, Alexandre M M; Pais de Barros, Jean-Paul; Arnould, Christine; Wanders, Ronald J; Trompier, Doriane; Savary, Stéphane

    2014-08-29

    ABCD1 and ABCD2 are two closely related ATP-binding cassette half-transporters predicted to homodimerize and form peroxisomal importers for fatty acyl-CoAs. Available evidence has shown that ABCD1 and ABCD2 display a distinct but overlapping substrate specificity, although much remains to be learned in this respect as well as in their capability to form functional heterodimers. Using a cell model expressing an ABCD2-EGFP fusion protein, we first demonstrated by proximity ligation assay and co-immunoprecipitation assay that ABCD1 interacts with ABCD2. Next, we tested in the pxa1/pxa2Δ yeast mutant the functionality of ABCD1/ABCD2 dimers by expressing chimeric proteins mimicking homo- or heterodimers. For further structure-function analysis of ABCD1/ABCD2 dimers, we expressed chimeric dimers fused to enhanced GFP in human skin fibroblasts of X-linked adrenoleukodystrophy patients. These cells are devoid of ABCD1 and accumulate very long-chain fatty acids (C26:0 and C26:1). We checked that the chimeric proteins were correctly expressed and targeted to the peroxisomes. Very long-chain fatty acid levels were partially restored in transfected X-linked adrenoleukodystrophy fibroblasts regardless of the chimeric construct used, thus demonstrating functionality of both homo- and heterodimers. Interestingly, the level of C24:6 n-3, the immediate precursor of docosahexaenoic acid, was decreased in cells expressing chimeric proteins containing at least one ABCD2 moiety. Our data demonstrate for the first time that both homo- and heterodimers of ABCD1 and ABCD2 are functionally active. Interestingly, the role of ABCD2 (in homo- and heterodimeric forms) in the metabolism of polyunsaturated fatty acids is clearly evidenced, and the chimeric dimers provide a novel tool to study substrate specificity of peroxisomal ATP-binding cassette transporters. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  9. Molecular cloning and functional characterization of an ATP-binding cassette transporter OtrC from Streptomyces rimosus

    Directory of Open Access Journals (Sweden)

    Yu Lan

    2012-08-01

    Full Text Available Abstract Background The otrC gene of Streptomyces rimosus was previously annotated as an oxytetracycline (OTC resistance protein. However, the amino acid sequence analysis of OtrC shows that it is a putative ATP-binding cassette (ABC transporter with multidrug resistance function. To our knowledge, none of the ABC transporters in S. rimosus have yet been characterized. In this study, we aimed to characterize the multidrug exporter function of OtrC and evaluate its relevancy to OTC production. Results In order to investigate OtrC’s function, otrC is cloned and expressed in E. coli The exporter function of OtrC was identified by ATPase activity determination and ethidium bromide efflux assays. Also, the susceptibilities of OtrC-overexpressing cells to several structurally unrelated drugs were compared with those of OtrC-non-expressing cells by minimal inhibitory concentration (MIC assays, indicating that OtrC functions as a drug exporter with a broad range of drug specificities. The OTC production was enhanced by 1.6-fold in M4018 (P = 0.000877 and 1.4-fold in SR16 (P = 0.00973 duplication mutants, while it decreased to 80% in disruption mutants (P = 0.0182 and 0.0124 in M4018 and SR16, respectively. Conclusions The results suggest that OtrC is an ABC transporter with multidrug resistance function, and plays an important role in self-protection by drug efflux mechanisms. This is the first report of such a protein in S. rimosus, and otrC could be a valuable target for genetic manipulation to improve the production of industrial antibiotics.

  10. Linsitinib (OSI-906) antagonizes ATP-binding cassette subfamily G member 2 and subfamily C member 10-mediated drug resistance.

    Science.gov (United States)

    Zhang, Hui; Kathawala, Rishil J; Wang, Yi-Jun; Zhang, Yun-Kai; Patel, Atish; Shukla, Suneet; Robey, Robert W; Talele, Tanaji T; Ashby, Charles R; Ambudkar, Suresh V; Bates, Susan E; Fu, Li-Wu; Chen, Zhe-Sheng

    2014-06-01

    In this study we investigated the effect of linsitinib on the reversal of multidrug resistance (MDR) mediated by the overexpression of the ATP-binding cassette (ABC) subfamily members ABCB1, ABCG2, ABCC1 and ABCC10. Our results indicate for the first time that linsitinib significantly potentiate the effect of anti-neoplastic drugs mitoxantrone (MX) and SN-38 in ABCG2-overexpressing cells; paclitaxel, docetaxel and vinblastine in ABCC10-overexpressing cells. Linsitinib moderately enhanced the cytotoxicity of vincristine in cell lines overexpressing ABCB1, whereas it did not alter the cytotoxicity of substrates of ABCC1. Furthermore, linsitinib significantly increased the intracellular accumulation and decreased the efflux of [(3)H]-MX in ABCG2-overexpressing cells and [(3)H]-paclitaxel in ABCC10-overexpressing cells. However, linsitinib, at a concentration that reversed MDR, did not significantly alter the expression levels of either the ABCG2 or ABCC10 transporter proteins. Furthermore, linsitinib did not significantly alter the intracellular localization of ABCG2 or ABCC10. Moreover, linsitinib stimulated the ATPase activity of ABCG2 in a concentration-dependent manner. Overall, our study suggests that linsitinib attenuates ABCG2- and ABCC10-mediated MDR by directly inhibiting their function as opposed to altering ABCG2 or ABCC10 protein expression. Published by Elsevier Ltd.

  11. Structural and functional characterization of an orphan ATP-binding cassette ATPase involved in manganese utilization and tolerance in Leptospira spp.

    Science.gov (United States)

    Benaroudj, Nadia; Saul, Frederick; Bellalou, Jacques; Miras, Isabelle; Weber, Patrick; Bondet, Vincent; Murray, Gerald L; Adler, Ben; Ristow, Paula; Louvel, Hélène; Haouz, Ahmed; Picardeau, Mathieu

    2013-12-01

    Pathogenic Leptospira species are the etiological agents of the widespread zoonotic disease leptospirosis. Most organisms, including Leptospira, require divalent cations for proper growth, but because of their high reactivity, these metals are toxic at high concentrations. Therefore, bacteria have acquired strategies to maintain metal homeostasis, such as metal import and efflux. By screening Leptospira biflexa transposon mutants for their ability to use Mn(2+), we have identified a gene encoding a putative orphan ATP-binding cassette (ABC) ATPase of unknown function. Inactivation of this gene in both L. biflexa and L. interrogans strains led to mutants unable to grow in medium in which iron was replaced by Mn(2+), suggesting an involvement of this ABC ATPase in divalent cation uptake. A mutation in this ATPase-coding gene increased susceptibility to Mn(2+) toxicity. Recombinant ABC ATPase of the pathogen L. interrogans exhibited Mg(2+)-dependent ATPase activity involving a P-loop motif. The structure of this ATPase was solved from a crystal containing two monomers in the asymmetric unit. Each monomer adopted a canonical two-subdomain organization of the ABC ATPase fold with an α/β subdomain containing the Walker motifs and an α subdomain containing the ABC signature motif (LSSGE). The two monomers were arranged in a head-to-tail orientation, forming a V-shaped particle with all the conserved ABC motifs at the dimer interface, similar to functional ABC ATPases. These results provide the first structural and functional characterization of a leptospiral ABC ATPase.

  12. Functional Analysis of an ATP-Binding Cassette Transporter Gene in Botrytis cinerea by Gene Disruption

    OpenAIRE

    Masami, NAKAJIMA; Junko, SUZUKI; Takehiko, HOSAKA; Tadaaki, HIBI; Katsumi, AKUTSU; School of Agriculture, Ibaraki University; School of Agriculture, Ibaraki University; School of Agriculture, Ibaraki University; Department of Agriculture and Environmental Biology, The University of Tokyo; School of Agriculture, Ibaraki University

    2001-01-01

    The BMR1 gene encoding an ABC transporter was cloned from Botrytis cinerea. To examine the function of BMR1 in B.cinerea, we isolated BMR1-deficient mutants after gene disruption. Disruption vector pBcDF4 was constructed by replacing the BMR1-coding region with a hygromycin B phosphotransferase gene(hph)cassette. The BMR1 disruptants had an increased sensitivity to polyoxin and iprobenfos. Polyoxin and iprobenfos, structurally unrelated compounds, may therefore be substrates of BMR1.

  13. LrABCF1, a GCN-type ATP-binding cassette transporter from Lilium regale, is involved in defense responses against viral and fungal pathogens.

    Science.gov (United States)

    Sun, Daoyang; Zhang, Xinguo; Li, Shaohua; Jiang, Cai-Zhong; Zhang, Yanlong; Niu, Lixin

    2016-12-01

    The L. regale ATP-binding cassette transporter gene, LrABCF1 belonging to GCN subfamily, functions as a positive regulator of plant defense against Cucumber mosaic virus, Tobacco rattle virus , and Botrytis cinerea in petunia. ATP-binding cassette (ABC) transporters are essential for membrane translocation in diverse biological processes, such as plant development and defense response. Here, a general control non-derepressible (GCN)-type ABC transporter gene, designated LrABCF1, was identified from Cucumber mosaic virus (CMV)-induced cDNA library of L. regale. LrABCF1 was up-regulated upon inoculation with CMV and Lily mottle virus (LMoV). Salicylic acid (SA) and ethylene (ET) application and treatments with abiotic stresses such as cold, high salinity, and wounding increased the transcript abundances of LrABCF1. Constitutive overexpression of LrABCF1 in petunia (Petunia × hybrida) resulted in an impairment of plant growth and development. LrABCF1 overexpression conferred reduced susceptibility to CMV, Tobacco rattle virus (TRV), and B. cinerea infection in transgenic petunia plants, accompanying by elevated transcripts of PhGCN2 and a few defense-related genes in SA-signaling pathway. Our data indicate that LrABCF1 positively modulates viral and fungal resistance.

  14. Hydrolysis at One of the Two Nucleotide-binding Sites Drives the Dissociation of ATP-binding Cassette Nucleotide-binding Domain Dimers*

    Science.gov (United States)

    Zoghbi, Maria E.; Altenberg, Guillermo A.

    2013-01-01

    The functional unit of ATP-binding cassette (ABC) transporters consists of two transmembrane domains and two nucleotide-binding domains (NBDs). ATP binding elicits association of the two NBDs, forming a dimer in a head-to-tail arrangement, with two nucleotides “sandwiched” at the dimer interface. Each of the two nucleotide-binding sites is formed by residues from the two NBDs. We recently found that the prototypical NBD MJ0796 from Methanocaldococcus jannaschii dimerizes in response to ATP binding and dissociates completely following ATP hydrolysis. However, it is still unknown whether dissociation of NBD dimers follows ATP hydrolysis at one or both nucleotide-binding sites. Here, we used luminescence resonance energy transfer to study heterodimers formed by one active (donor-labeled) and one catalytically defective (acceptor-labeled) NBD. Rapid mixing experiments in a stop-flow chamber showed that NBD heterodimers with one functional and one inactive site dissociated at a rate indistinguishable from that of dimers with two hydrolysis-competent sites. Comparison of the rates of NBD dimer dissociation and ATP hydrolysis indicated that dissociation followed hydrolysis of one ATP. We conclude that ATP hydrolysis at one nucleotide-binding site drives NBD dimer dissociation. PMID:24129575

  15. Overexpression of the ATP-binding cassette half-transporter, ABCG2 (Mxr/BCrp/ABCP1), in flavopiridol-resistant human breast cancer cells

    DEFF Research Database (Denmark)

    Robey, R W; Medina-Pérez, W Y; Nishiyama, K

    2001-01-01

    We sought to characterize the interactions of flavopiridol with members of the ATP-binding cassette (ABC) transporter family. Cells overexpressing multidrug resistance-1 (MDR-1) and multidrug resistance-associated protein (MRP) did not exhibit appreciable flavopiridol resistance, whereas cell lines...... overexpressing the ABC half-transporter, ABCG2 (MXR/BCRP/ABCP1), were found to be resistant to flavopiridol. Flavopiridol at a concentration of 10 microM was able to prevent MRP-mediated calcein efflux, whereas Pgp-mediated transport of rhodamine 123 was unaffected at flavopiridol concentrations of up to 100...... microM. To determine putative mechanisms of resistance to flavopiridol, we exposed the human breast cancer cell line MCF-7 to incrementally increasing concentrations of flavopiridol. The resulting resistant subline, MCF-7 FLV1000, is maintained in 1,000 nM flavopiridol and was found to be 24-fold...

  16. Overexpression of the ATP-binding cassette half-transporter, ABCG2 (Mxr/BCrp/ABCP1), in flavopiridol-resistant human breast cancer cells

    DEFF Research Database (Denmark)

    Robey, R W; Medina-Pérez, W Y; Nishiyama, K

    2001-01-01

    We sought to characterize the interactions of flavopiridol with members of the ATP-binding cassette (ABC) transporter family. Cells overexpressing multidrug resistance-1 (MDR-1) and multidrug resistance-associated protein (MRP) did not exhibit appreciable flavopiridol resistance, whereas cell lines...... overexpressing the ABC half-transporter, ABCG2 (MXR/BCRP/ABCP1), were found to be resistant to flavopiridol. Flavopiridol at a concentration of 10 microM was able to prevent MRP-mediated calcein efflux, whereas Pgp-mediated transport of rhodamine 123 was unaffected at flavopiridol concentrations of up to 100...... resistant to flavopiridol, as well as highly cross-resistant to mitoxantrone (675-fold), topotecan (423-fold), and SN-38 (950-fold), the active metabolite of irinotecan. Because this cross-resistance pattern is consistent with that reported for ABCG2-overexpressing cells, cytotoxicity studies were repeated...

  17. A conserved mitochondrial ATP-binding cassette transporter exports glutathione polysulfide for cytosolic metal cofactor assembly.

    Science.gov (United States)

    Schaedler, Theresia A; Thornton, Jeremy D; Kruse, Inga; Schwarzländer, Markus; Meyer, Andreas J; van Veen, Hendrik W; Balk, Janneke

    2014-08-22

    An ATP-binding cassette transporter located in the inner mitochondrial membrane is involved in iron-sulfur cluster and molybdenum cofactor assembly in the cytosol, but the transported substrate is unknown. ATM3 (ABCB25) from Arabidopsis thaliana and its functional orthologue Atm1 from Saccharomyces cerevisiae were expressed in Lactococcus lactis and studied in inside-out membrane vesicles and in purified form. Both proteins selectively transported glutathione disulfide (GSSG) but not reduced glutathione in agreement with a 3-fold stimulation of ATPase activity by GSSG. By contrast, Fe(2+) alone or in combination with glutathione did not stimulate ATPase activity. Arabidopsis atm3 mutants were hypersensitive to an inhibitor of glutathione biosynthesis and accumulated GSSG in the mitochondria. The growth phenotype of atm3-1 was strongly enhanced by depletion of the mitochondrion-localized, GSH-dependent persulfide oxygenase ETHE1, suggesting that the physiological substrate of ATM3 contains persulfide in addition to glutathione. Consistent with this idea, a transportomics approach using mass spectrometry showed that glutathione trisulfide (GS-S-SG) was transported by Atm1. We propose that mitochondria export glutathione polysulfide, containing glutathione and persulfide, for iron-sulfur cluster assembly in the cytosol. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  18. Molecular phylogenetic study and expression analysis of ATP-binding cassette transporter gene family in Oryza sativa in response to salt stress.

    Science.gov (United States)

    Saha, Jayita; Sengupta, Atreyee; Gupta, Kamala; Gupta, Bhaskar

    2015-02-01

    ATP-binding cassette (ABC) transporter is a large gene superfamily that utilizes the energy released from ATP hydrolysis for transporting myriad of substrates across the biological membranes. Although many investigations have been done on the structural and functional analysis of the ABC transporters in Oryza sativa, much less is known about molecular phylogenetic and global expression pattern of the complete ABC family in rice. In this study, we have carried out a comprehensive phylogenetic analysis constructing neighbor-joining and maximum-likelihood trees based on various statistical methods of different ABC protein subfamily of five plant lineages including Chlamydomonas reinhardtii (green algae), Physcomitrella patens (moss), Selaginella moellendorffii (lycophyte), Arabidopsis thaliana (dicot) and O. sativa (monocot) to explore the origin and evolutionary patterns of these ABC genes. We have identified several conserved motifs in nucleotide binding domain (NBD) of ABC proteins among all plant lineages during evolution. Amongst the different ABC protein subfamilies, 'ABCE' has not yet been identified in lower plant genomes (algae, moss and lycophytes). The result indicated that gene duplication and diversification process acted upon these genes as a major operative force creating new groups and subgroups and functional divergence during evolution. We have demonstrated that rice ABCI subfamily consists of only half size transporters that represented highly dynamic members showing maximum sequence variations among the other rice ABC subfamilies. The evolutionary and the expression analysis contribute to a deep insight into the evolution and diversity of rice ABC proteins and their roles in response to salt stress that facilitate our further understanding on rice ABC transporters. Copyright © 2014 Elsevier Ltd. All rights reserved.

  19. Endothelial ATP-binding cassette G1 in mouse endothelium protects against hemodynamic-induced atherosclerosis

    International Nuclear Information System (INIS)

    Xue, Shanshan; Wang, Jiaxing; Zhang, Xu; Shi, Ying; Li, Bochuan; Bao, Qiankun; Pang, Wei; Ai, Ding; Zhu, Yi; He, Jinlong

    2016-01-01

    Activated vascular endothelium inflammation under persistent hyperlipidemia is the initial step of atherogenesis. ATP-binding cassette G1 (ABCG1) is a crucial factor maintaining sterol and lipid homeostasis by transporting cholesterol efflux to high-density lipoprotein. In this study, we investigated the protective effects of ABCG1 in endothelial inflammation activation during early-stage atherogenesis in mice and the underlying mechanisms. Endothelial cell (EC)-specific ABCG1 transgenic (EC-ABCG1-Tg) mice were generated and cross-bred with low-density lipoprotein receptor–deficient (Ldlr −/− ) mice. After a 4-week Western-type diet, the mice were sacrificed for assessing atherosclerosis. Human umbilical vein ECs were treated with different flows, and ABCG1 was adenovirally overexpressed to investigate the mechanism in vitro. Compared with Ldlr −/− mouse aortas, EC-ABCG1-Tg/Ldlr −/− aortas showed decreased early-stage lesions. Furthermore, the lesion area in the EC-ABCG1-Tg/Ldlr −/− mouse aortic arch but not thoracic aorta was significantly reduced, which suggests a protective role of ABCG1 under atheroprone flow. In vitro, overexpression of ABCG1 attenuated EC activation caused by oscillatory shear stress. Overexpression of ABCG1 blunted cholesterol-activated ECs in vitro. In exploring the mechanisms of ABCG1 attenuating endothelial inflammation, we found that ABCG1 inhibited oscillatory flow-activated nuclear factor kappa B and NLRP3 inflammasome in ECs. ABCG1 may play a protective role in early-stage atherosclerosis by reducing endothelial activation induced by oscillatory shear stress via suppressing the inflammatory response. - Highlights: • EC-ABCG1-Tg mice in a Ldlr −/− background showed decreased atherosclerosis. • Overexpression of ABCG1 in ECs decreased OSS-induced EC activation. • NLRP3 and NF-κB might be an underlying mechanism of ABCG1 protective role.

  20. A novel flow cytometric HTS assay reveals functional modulators of ATP binding cassette transporter ABCB6.

    Directory of Open Access Journals (Sweden)

    Kishore Polireddy

    Full Text Available ABCB6 is a member of the adenosine triphosphate (ATP-binding cassette family of transporter proteins that is increasingly recognized as a relevant physiological and therapeutic target. Evaluation of modulators of ABCB6 activity would pave the way toward a more complete understanding of the significance of this transport process in tumor cell growth, proliferation and therapy-related drug resistance. In addition, this effort would improve our understanding of the function of ABCB6 in normal physiology with respect to heme biosynthesis, and cellular adaptation to metabolic demand and stress responses. To search for modulators of ABCB6, we developed a novel cell-based approach that, in combination with flow cytometric high-throughput screening (HTS, can be used to identify functional modulators of ABCB6. Accumulation of protoporphyrin, a fluorescent molecule, in wild-type ABCB6 expressing K562 cells, forms the basis of the HTS assay. Screening the Prestwick Chemical Library employing the HTS assay identified four compounds, benzethonium chloride, verteporfin, tomatine hydrochloride and piperlongumine, that reduced ABCB6 mediated cellular porphyrin levels. Validation of the identified compounds employing the hemin-agarose affinity chromatography and mitochondrial transport assays demonstrated that three out of the four compounds were capable of inhibiting ABCB6 mediated hemin transport into isolated mitochondria. However, only verteporfin and tomatine hydrochloride inhibited ABCB6's ability to compete with hemin as an ABCB6 substrate. This assay is therefore sensitive, robust, and suitable for automation in a high-throughput environment as demonstrated by our identification of selective functional modulators of ABCB6. Application of this assay to other libraries of synthetic compounds and natural products is expected to identify novel modulators of ABCB6 activity.

  1. Molecular Events Involved in a Single Cycle of Ligand Transfer from an ATP Binding Cassette Transporter, LolCDE, to a Molecular Chaperone, LolA*

    OpenAIRE

    Taniguchi, Naohiro; Tokuda, Hajime

    2008-01-01

    An ATP binding cassette transporter LolCDE complex releases lipoproteins from the inner membrane of Escherichia coli in an ATP-dependent manner, leading to the formation of a complex between a lipoprotein and a periplasmic chaperone, LolA. LolA is proposed to undergo a conformational change upon the lipoprotein binding. The lipoprotein is then transferred from the LolA-lipoprotein complex to the outer membrane via LolB. Unlike most ATP binding cassette transporters med...

  2. Role of ATP-binding cassette and solute carrier transporters in erlotinib CNS penetration and intracellular accumulation.

    Science.gov (United States)

    Elmeliegy, Mohamed A; Carcaboso, Angel M; Tagen, Michael; Bai, Feng; Stewart, Clinton F

    2011-01-01

    To study the role of drug transporters in central nervous system (CNS) penetration and cellular accumulation of erlotinib and its metabolite, OSI-420. After oral erlotinib administration to wild-type and ATP-binding cassette (ABC) transporter-knockout mice (Mdr1a/b(-/-), Abcg2(-/-), Mdr1a/b(-/-)Abcg2(-/-), and Abcc4(-/-)), plasma was collected and brain extracellular fluid (ECF) was sampled using intracerebral microdialysis. A pharmacokinetic model was fit to erlotinib and OSI-420 concentration-time data, and brain penetration (P(Brain)) was estimated by the ratio of ECF-to-unbound plasma area under concentration-time curves. Intracellular accumulation of erlotinib was assessed in cells overexpressing human ABC transporters or SLC22A solute carriers. P(Brain) in wild-type mice was 0.27 ± 0.11 and 0.07 ± 0.02 (mean ± SD) for erlotinib and OSI-420, respectively. Erlotinib and OSI-420 P(Brain) in Abcg2(-/-) and Mdr1a/b(-/-)Abcg2(-/-) mice were significantly higher than in wild-type mice. Mdr1a/b(-/-) mice showed similar brain ECF penetration as wild-type mice (0.49 ± 0.37 and 0.04 ± 0.02 for erlotinib and OSI-420, respectively). In vitro, erlotinib and OSI-420 accumulation was significantly lower in cells overexpressing breast cancer resistance protein (BCRP) than in control cells. Only OSI-420, not erlotinib, showed lower accumulation in cells overexpressing P-glycoprotein (P-gp) than in control cells. The P-gp/BCRP inhibitor elacridar increased erlotinib and OSI-420 accumulation in BCRP-overexpressing cells. Erlotinib uptake was higher in OAT3- and OCT2-transfected cells than in empty vector control cells. Abcg2 is the main efflux transporter preventing erlotinib and OSI-420 penetration in mouse brain. Erlotinib and OSI-420 are substrates for SLC22A family members OAT3 and OCT2. Our findings provide a mechanistic basis for erlotinib CNS penetration, cellular uptake, and efflux mechanisms. ©2010 AACR.

  3. Endothelial ATP-binding cassette G1 in mouse endothelium protects against hemodynamic-induced atherosclerosis

    Energy Technology Data Exchange (ETDEWEB)

    Xue, Shanshan [Department of Physiology and Pathophysiology, Tianjin Medical University, Tianjin, 300070 (China); Department of Pediatrics, Baodi District People’s Hospital of Tianjin City, Tianjin, 301800 (China); Wang, Jiaxing [Department of Physiology and Pathophysiology, Peking University Health Science Center, Beijing, 100191 (China); Zhang, Xu; Shi, Ying; Li, Bochuan; Bao, Qiankun [Department of Physiology and Pathophysiology, Tianjin Medical University, Tianjin, 300070 (China); Pang, Wei [Department of Physiology and Pathophysiology, Peking University Health Science Center, Beijing, 100191 (China); Ai, Ding [Department of Physiology and Pathophysiology, Tianjin Medical University, Tianjin, 300070 (China); Zhu, Yi [Department of Physiology and Pathophysiology, Tianjin Medical University, Tianjin, 300070 (China); Department of Physiology and Pathophysiology, Peking University Health Science Center, Beijing, 100191 (China); He, Jinlong, E-mail: hejinlong@tmu.edu.cn [Department of Physiology and Pathophysiology, Tianjin Medical University, Tianjin, 300070 (China)

    2016-08-19

    Activated vascular endothelium inflammation under persistent hyperlipidemia is the initial step of atherogenesis. ATP-binding cassette G1 (ABCG1) is a crucial factor maintaining sterol and lipid homeostasis by transporting cholesterol efflux to high-density lipoprotein. In this study, we investigated the protective effects of ABCG1 in endothelial inflammation activation during early-stage atherogenesis in mice and the underlying mechanisms. Endothelial cell (EC)-specific ABCG1 transgenic (EC-ABCG1-Tg) mice were generated and cross-bred with low-density lipoprotein receptor–deficient (Ldlr{sup −/−}) mice. After a 4-week Western-type diet, the mice were sacrificed for assessing atherosclerosis. Human umbilical vein ECs were treated with different flows, and ABCG1 was adenovirally overexpressed to investigate the mechanism in vitro. Compared with Ldlr{sup −/−} mouse aortas, EC-ABCG1-Tg/Ldlr{sup −/−} aortas showed decreased early-stage lesions. Furthermore, the lesion area in the EC-ABCG1-Tg/Ldlr{sup −/−} mouse aortic arch but not thoracic aorta was significantly reduced, which suggests a protective role of ABCG1 under atheroprone flow. In vitro, overexpression of ABCG1 attenuated EC activation caused by oscillatory shear stress. Overexpression of ABCG1 blunted cholesterol-activated ECs in vitro. In exploring the mechanisms of ABCG1 attenuating endothelial inflammation, we found that ABCG1 inhibited oscillatory flow-activated nuclear factor kappa B and NLRP3 inflammasome in ECs. ABCG1 may play a protective role in early-stage atherosclerosis by reducing endothelial activation induced by oscillatory shear stress via suppressing the inflammatory response. - Highlights: • EC-ABCG1-Tg mice in a Ldlr{sup −/−} background showed decreased atherosclerosis. • Overexpression of ABCG1 in ECs decreased OSS-induced EC activation. • NLRP3 and NF-κB might be an underlying mechanism of ABCG1 protective role.

  4. Analysis of the structural and functional roles of coupling helices in the ATP-binding cassette transporter MsbA through enzyme assays and molecular dynamics simulations.

    Science.gov (United States)

    Furuta, Tadaomi; Yamaguchi, Tomohiro; Kato, Hiroaki; Sakurai, Minoru

    2014-07-08

    ATP-binding cassette (ABC) transporters are constructed from some common structural units: the highly conserved nucleotide-binding domains (NBDs), which work as a nucleotide-dependent engine for driving substrate transport, the diverse transmembrane domains (TMDs), which create the translocation pathway, and the coupling helices (CHs), which are located at the NBD-TMD interface. Although the CHs are believed to be essential for NBD-TMD communication, their roles remain unclear. In this study, we performed enzyme assays and molecular dynamics (MD) simulations of the ABC transporter MsbA and two MsbA mutants in which the amino acid residues of one of the CHs were mutated to alanines: (i) wild type (Wt), (ii) CH1 mutant (Mt1), and (iii) CH2 mutant (Mt2). The experiments show that the CH2 mutation decreases the ATPase activity (kcat) compared with that of the Wt (a decrease of 32%), and a nearly equal degree of decrease in the ATP binding affinity (Km) was observed for both Mt1 and Mt2. The MD simulations successfully accounted for several structural and dynamical origins for these experimental observations. In addition, on the basis of collective motion and morphing analyses, we propose that the reverse-rotational motions and noddinglike motions between the NBDs and TMDs are indispensable for the conformational transition between the inward- and outward-facing conformations. In particular, CH2 is significantly important for the occurrence of the noddinglike motion. These findings provide important insights into the structure-function relationship of ABC transporters.

  5. ATP-binding cassette-like transporters are involved in the transport of lignin precursors across plasma and vacuolar membranes

    Energy Technology Data Exchange (ETDEWEB)

    Miao, Y.C.; Liu, C.

    2010-12-28

    Lignin is a complex biopolymer derived primarily from the condensation of three monomeric precursors, the monolignols. The synthesis of monolignols occurs in the cytoplasm. To reach the cell wall where they are oxidized and polymerized, they must be transported across the cell membrane. However, the molecular mechanisms underlying the transport process are unclear. There are conflicting views about whether the transport of these precursors occurs by passive diffusion or is an energized active process; further, we know little about what chemical forms are required. Using isolated plasma and vacuolar membrane vesicles prepared from Arabidopsis, together with applying different transporter inhibitors in the assays, we examined the uptake of monolignols and their derivatives by these native membrane vesicles. We demonstrate that the transport of lignin precursors across plasmalemma and their sequestration into vacuoles are ATP-dependent primary-transport processes, involving ATP-binding cassette-like transporters. Moreover, we show that both plasma and vacuolar membrane vesicles selectively transport different forms of lignin precursors. In the presence of ATP, the inverted plasma membrane vesicles preferentially take up monolignol aglycones, whereas the vacuolar vesicles are more specific for glucoconjugates, suggesting that the different ATP-binding cassette-like transporters recognize different chemical forms in conveying them to distinct sites, and that glucosylation of monolignols is necessary for their vacuolar storage but not required for direct transport into the cell wall in Arabidopsis.

  6. Conformational changes of the bacterial type I ATP-binding cassette importer HisQMP2 at distinct steps of the catalytic cycle.

    Science.gov (United States)

    Heuveling, Johanna; Frochaux, Violette; Ziomkowska, Joanna; Wawrzinek, Robert; Wessig, Pablo; Herrmann, Andreas; Schneider, Erwin

    2014-01-01

    Prokaryotic solute binding protein-dependent ATP-binding cassette import systems are divided into type I and type II and mechanistic differences in the transport process going along with this classification are under intensive investigation. Little is known about the conformational dynamics during the catalytic cycle especially concerning the transmembrane domains. The type I transporter for positively charged amino acids from Salmonella enterica serovar Typhimurium (LAO-HisQMP2) was studied by limited proteolysis in detergent solution in the absence and presence of co-factors including ATP, ADP, LAO/arginine, and Mg(2+) ions. Stable peptide fragments could be obtained and differentially susceptible cleavage sites were determined by mass spectrometry as Lys-258 in the nucleotide-binding subunit, HisP, and Arg-217/Arg-218 in the transmembrane subunit, HisQ. In contrast, transmembrane subunit HisM was gradually degraded but no stable fragment could be detected. HisP and HisQ were equally resistant under pre- and post-hydrolysis conditions in the presence of arginine-loaded solute-binding protein LAO and ATP/ADP. Some protection was also observed with LAO/arginine alone, thus reflecting binding to the transporter in the apo-state and transmembrane signaling. Comparable digestion patterns were obtained with the transporter reconstituted into proteoliposomes and nanodiscs. Fluorescence lifetime spectroscopy confirmed the change of HisQ(R218) to a more apolar microenvironment upon ATP binding and hydrolysis. Limited proteolysis was subsequently used as a tool to study the consequences of mutations on the transport cycle. Together, our data suggest similar conformational changes during the transport cycle as described for the maltose ABC transporter of Escherichia coli, despite distinct structural differences between both systems. © 2013.

  7. AtMRP2, an Arabidopsis ATP binding cassette transporter able to transport glutathione S-conjugates and chlorophyll catabolites: functional comparisons with Atmrp1.

    Science.gov (United States)

    Lu, Y P; Li, Z S; Drozdowicz, Y M; Hortensteiner, S; Martinoia, E; Rea, P A

    1998-02-01

    Three ATP binding cassette (ABC) transporter-like activities directed toward large amphipathic organic anions have recently been identified on the vacuolar membrane of plant cells. These are the Mg-ATP-energized, vanadate-inhibitable vacuolar accumulation of glutathione S-conjugates (GS conjugates), chlorophyll catabolites, and bile acids, respectively. Although each of these activities previously had been assigned to distinct pumps in native plant membranes, we describe here the molecular cloning, physical mapping, and heterologous expression of a gene, AtMRP2, from Arabidopsis thaliana that encodes a multispecific ABC transporter competent in the transport of both GS conjugates and chlorophyll catabolites. Unlike its isoform, AtMRP1, which transports the model Brassica napus chlorophyll catabolite transporter substrate Bn-NCC-1 at low efficiency, heterologously expressed AtMRP2 has the facility for simultaneous high-efficiency parallel transport of GS conjugates and Bn-NCC-1. The properties of AtMRP2 therefore establish a basis for the manipulation of two previously identified plant ABC transporter activities and provide an explanation for how the comparable transporter in native plant membranes would be systematically mistaken for two distinct transporters. These findings are discussed with respect to the functional organization of AtMRP2, the inability of AtMRP2 and AtMRP1 to transport the model bile acid transporter substrate taurocholate (despite the pronounced sensitivity of both to direct inhibition by this agent), the differential patterns of expression of their genes in the intact plant, and the high capacity of AtMRP2 for the transport of glutathionated herbicides and anthocyanins.

  8. The capacity of Listeria monocytogenes mutants with in-frame deletions in putative ATP-binding cassette transporters to form biofilms and comparison with the wild type

    Directory of Open Access Journals (Sweden)

    Marina Ceruso

    2014-02-01

    Full Text Available Listeria monocytogenes (Lm is a food-borne pathogen responsible for human listeriosis, an invasive infection with high mortality rates. Lm has developed efficient strategies for survival under stress conditions such as starvation and wide variations in temperature, pH, and osmolarity. Therefore, Lm can survive in food under multiple stress conditions. Detailed studies to determine the mode of action of this pathogen for survival under stress conditions are important to control Lm in food. It has been shown that genes encoding for ATP-binding cassette (ABC transporters are induced in Lm in food, in particular under stress conditions. Previous studies showed that these genes are involved in sensitivity to nisin, acids, and salt. The aim of this study was to determine the involvement of some ABC transporters in biofilm formation. Therefore, deletion mutants of ABC transporter genes (LMOf2365_1875 and LMOf2365_1877 were created in Lm F2365, and then were compared to the wild type for their capacity to form biofilms. Lm strain F2365 was chosen as reference since the genome is fully sequenced and furthermore this strain is particularly involved in food-borne outbreaks of listeriosis. Our results showed that DLMOf2365_1875 had an increased capacity to form biofilms compared to the wild type, indicating that LMOf2365_1875 negatively regulates biofilm formation. A deeper knowledge on the ability to form biofilms in these mutants may help in the development of intervention strategies to control Lm in food and in the environment.

  9. Oct-3/4 modulates the drug-resistant phenotype of glioblastoma cells through expression of ATP binding cassette transporter G2.

    Science.gov (United States)

    Hosokawa, Yuki; Takahashi, Hisaaki; Inoue, Akihiro; Kawabe, Yuya; Funahashi, Yu; Kameda, Kenji; Sugimoto, Kana; Yano, Hajime; Harada, Hironobu; Kohno, Shohei; Ohue, Shiro; Ohnishi, Takanori; Tanaka, Junya

    2015-06-01

    Drug resistance is a major obstacle for the efficacy of chemotherapeutic treatment of tumors. Oct-3/4, a self-renewal regulator in stem cells, is expressed in various kinds of solid tumors including glioblastoma. Although Oct-3/4 expression has been implicated in the malignancy and prognosis of glioblastomas, little is known of its involvement in drug resistances of glioblastoma. The involvement of Oct-3/4 in drug resistance of glioblastoma cells was assessed by lactate dehydrogenase assay, efflux assay of an anticancer drug, poly ADP-ribose polymerase cleavage, and in vivo xenograft experiments. Involvement of a drug efflux pump ATP binding cassette transporter G2 in Oct-3/4-induced drug resistance was evaluated by quantitative PCR analysis and knockdown by shRNA. Oct-3/4 decreased the susceptibility to chemotherapeutic drugs by enhancing excretion of drugs through a drug efflux pump gene, ATP binding cassette transporter G2. Moreover, the expression of Oct-3/4 was well correlated to ATP binding cassette transporter G2 expression in clinical GB tissues. Oct-3/4 elevated the ATP binding cassette transporter G2 expression, leading to acquisition of a drug-resistant phenotype by glioblastoma cells. If the drug-resistance of glioblastoma cells could be suppressed, it should be a highly ameliorative treatment for glioblastoma patients. Therefore, signaling pathways from Oct-3/4 to ATP binding cassette transporter G2 should be intensively elucidated to develop new therapeutic interventions for better efficacy of anti-cancer drugs. Copyright © 2015 Elsevier B.V. All rights reserved.

  10. Role of NH{sub 2}-terminal hydrophobic motif in the subcellular localization of ATP-binding cassette protein subfamily D: Common features in eukaryotic organisms

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Asaka; Asahina, Kota; Okamoto, Takumi; Kawaguchi, Kosuke [Department of Biological Chemistry, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, 2630 Sugitani, Toyama 930-0194 (Japan); Kostsin, Dzmitry G. [Department of Biological Chemistry, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, 2630 Sugitani, Toyama 930-0194 (Japan); Institute of Biophysics and Cell Engineering, National Academy of Sciences of Belarus, Academicheskaya Str. 27, Minsk 220072 (Belarus); Kashiwayama, Yoshinori [Department of Biological Chemistry, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, 2630 Sugitani, Toyama 930-0194 (Japan); Takanashi, Kojiro; Yazaki, Kazufumi [Laboratory of Plant Gene Expression, Research Institute for Sustainable Humanosphere, Kyoko University, Uji, Kyoto 611-0011 (Japan); Imanaka, Tsuneo, E-mail: imanaka@pha.u-toyama.ac.jp [Department of Biological Chemistry, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, 2630 Sugitani, Toyama 930-0194 (Japan); Morita, Masashi [Department of Biological Chemistry, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, 2630 Sugitani, Toyama 930-0194 (Japan)

    2014-10-24

    Highlights: • ABCD proteins classifies based on with or without NH{sub 2}-terminal hydrophobic segment. • The ABCD proteins with the segment are targeted peroxisomes. • The ABCD proteins without the segment are targeted to the endoplasmic reticulum. • The role of the segment in organelle targeting is conserved in eukaryotic organisms. - Abstract: In mammals, four ATP-binding cassette (ABC) proteins belonging to subfamily D have been identified. ABCD1–3 possesses the NH{sub 2}-terminal hydrophobic region and are targeted to peroxisomes, while ABCD4 lacking the region is targeted to the endoplasmic reticulum (ER). Based on hydropathy plot analysis, we found that several eukaryotes have ABCD protein homologs lacking the NH{sub 2}-terminal hydrophobic segment (H0 motif). To investigate whether the role of the NH{sub 2}-terminal H0 motif in subcellular localization is conserved across species, we expressed ABCD proteins from several species (metazoan, plant and fungi) in fusion with GFP in CHO cells and examined their subcellular localization. ABCD proteins possessing the NH{sub 2}-terminal H0 motif were localized to peroxisomes, while ABCD proteins lacking this region lost this capacity. In addition, the deletion of the NH{sub 2}-terminal H0 motif of ABCD protein resulted in their localization to the ER. These results suggest that the role of the NH{sub 2}-terminal H0 motif in organelle targeting is widely conserved in living organisms.

  11. An ATP-binding cassette subfamily G full transporter is essential for the retention of leaf water in both wild barley and rice.

    Science.gov (United States)

    Chen, Guoxiong; Komatsuda, Takao; Ma, Jian Feng; Nawrath, Christiane; Pourkheirandish, Mohammad; Tagiri, Akemi; Hu, Yin-Gang; Sameri, Mohammad; Li, Xinrong; Zhao, Xin; Liu, Yubing; Li, Chao; Ma, Xiaoying; Wang, Aidong; Nair, Sudha; Wang, Ning; Miyao, Akio; Sakuma, Shun; Yamaji, Naoki; Zheng, Xiuting; Nevo, Eviatar

    2011-07-26

    Land plants have developed a cuticle preventing uncontrolled water loss. Here we report that an ATP-binding cassette (ABC) subfamily G (ABCG) full transporter is required for leaf water conservation in both wild barley and rice. A spontaneous mutation, eibi1.b, in wild barley has a low capacity to retain leaf water, a phenotype associated with reduced cutin deposition and a thin cuticle. Map-based cloning revealed that Eibi1 encodes an HvABCG31 full transporter. The gene was highly expressed in the elongation zone of a growing leaf (the site of cutin synthesis), and its gene product also was localized in developing, but not in mature tissue. A de novo wild barley mutant named "eibi1.c," along with two transposon insertion lines of rice mutated in the ortholog of HvABCG31 also were unable to restrict water loss from detached leaves. HvABCG31 is hypothesized to function as a transporter involved in cutin formation. Homologs of HvABCG31 were found in green algae, moss, and lycopods, indicating that this full transporter is highly conserved in the evolution of land plants.

  12. The role of ATP-binding cassette transporter genes in the progression of prostate cancer.

    Science.gov (United States)

    Karatas, Omer F; Guzel, Esra; Duz, Mehmet B; Ittmann, Michael; Ozen, Mustafa

    2016-04-01

    Prostate cancer (PCa) is the most commonly diagnosed neoplasm and the second leading cause of cancer-related death among men in developed countries. There is no clear evidence showing the success of current screening tests in reducing mortality of PCa. In this study, we aimed to profile expressions of nine ABC transporters, ABCA5, ABCB1, ABCB6, ABCC1, ABCC2, ABCC3, ABCC5, ABCC10, and ABCF2, in recurrent, non-recurrent PCa and normal prostate tissues. A total of 77 (39 recurrent, 38 non-recurrent) radical prostatectomy and 20 normal prostate samples, obtained from Baylor College of Medicine Prostate Cancer program, were included into the study and divided into two independent groups as test and validation sample sets. Differential expression of selected ABC transporters was assessed using quantitative real-time PCR (qRT-PCR). Pearson's correlation test, receiver operating characteristics (ROC) analysis and Kaplan-Meier test were used for statistical analysis. QRT-PCR results demonstrated the elevated expression of ABCA5, ABCB1, ABCB6, ABCC1, and ABCC2 as well as reduced expression of ABCC3 in PCa samples compared to normal prostate tissues. In addition, we found deregulation of ABCB1, ABCB6, ABCC3, and ABCC10 in recurrent PCa samples and validated differential expression of ABCB6, ABCC3, and ABCC10 in recurrent PCa compared to non-recurrent PCa. Pearson's correlation, ROC and Kaplan-Meier analysis revealed the power of these three ABC transporters for estimating prognosis of PCa. We demonstrated differential expression of ABC transporters both in tumor versus normal and recurrent versus non-recurrent comparisons. Our data suggest ABCB6, ABCC3, and ABCC10 as valuable predictors of PCa progression. © 2015 Wiley Periodicals, Inc.

  13. ATP-binding and -hydrolysis activities of ALDP (ABCD1) and ALDRP (ABCD2), human peroxisomal ABC proteins, overexpressed in Sf21 cells.

    Science.gov (United States)

    Morita, Masashi; Kurisu, Mikinori; Kashiwayama, Yoshinori; Yokota, Sadaki; Imanaka, Tsuneo

    2006-09-01

    The peroxisomal ATP-binding cassette (ABC) proteins, adrenoleukodystrophy protein (ALDP, ABCD1) and ALD-related protein (ALDRP, ABCD2), were expressed in Spodoptera frugiperda 21 (Sf21) insect cells using a baculovirus-mediated expression system. Immunoelectron microscopy and subcellular fractionation revealed that the overexpressed ALDP was distributed in various subcellular organelles including mitochondria, nucleus and peroxisomes. The ALDP was not extractable with Na(2)CO(3) treatment, suggesting that it integrated into membranes. ATPase activity was detected in the membrane fraction expressing ALDP. The nucleotide-binding capacities of the expressed ALDP were estimated by the binding to ATP- or ADP-agarose. ALDP exhibited an affinity to both ADP and ATP. In contrast, ALDRP exhibited an affinity to ADP but scarcely to ATP. The ALDP in the Sf21 membrane fraction was extracted with n-dodecyl-beta-maltoside and successively purified with a chelate column. The nucleotide-binding and ATPase activities of the purified ALDP were, however, not detected. It may be that certain membranous components are required for the activity. We demonstrate for the first time that the peroxisomal ABC proteins can be expressed in Sf21 membranes maintaining their nucleotide-binding abilities and ATPase activities, and the expressed proteins will be of use for further characterization.

  14. High resistance of Isaria fumosorosea to carbendazim arises from the overexpression of an ATP-binding cassette transporter (ifT1) rather than tubulin mutation.

    Science.gov (United States)

    Song, T-T; Ying, S-H; Feng, M-G

    2012-01-01

    Probing possible mechanisms involved in the resistance of entomopathogenic fungus Isaria fumosorosea to carbendazim fungicide. A carbendazim-sensitive strain (If116) selected from 15 wild-type strains was subjected to NaNO(2) -induced mutagenesis, yielding nine mutants with carbendazim resistance increased by 82- to 830-fold and thermotolerance decreased by 15-51%. Comparing the protein sequences deduced from the α- and β-tubulin genes of If116 and its mutants revealed no traceable site mutation relating to the enhanced resistance although the transcripts levels of β-tubulin gene in all mutants were 0·87- to 7·16-fold of that in If116. Three examined mutants showed multidrug resistance because they were significantly more resistant to glufosinate, imidacloprid and other six fungicides than If116 during growth. Further examination of rhodamine-stained blastospores revealed existence of drug efflux pump protein(s) in all carbendazim-resistant mutants. Thus, the sequences of an ATP-binding cassette (ABC) transporter gene (ifT1) and its promoter region cloned from the wild-type and mutant strains were analysed. Three common point mutations were located, respectively, at the binding sites of Gal4, Abf1 and Raf, which are crucial transcription factors in the regulative network of numerous protein loci. Such point mutations elevated the ifT1 expression by 17 to 137-fold in all the mutants. The overexpression of the ABC transporter caused by the point mutations at the binding sites was responsible for the fungal resistance to various pesticides including carbendazim. The transporter-mediated multidrug resistance found for the first time in entomopathogenic fungi is potential for use in improving mycoinsecticide compatibility with chemical pesticides. © 2011 The Authors. Journal of Applied Microbiology © 2011 The Society for Applied Microbiology.

  15. Functional roles of YPT31 and YPT32 in clotrimazole resistance of Saccharomyces cerevisiae through effects on vacuoles and ATP-binding cassette transporter(s).

    Science.gov (United States)

    Tsujimoto, Yoshiyuki; Takase, Daisuke; Okano, Hajime; Tomari, Naohiro; Watanabe, Kunihiko; Matsui, Hiroshi

    2013-01-01

    We identified YPT31, which is involved in Golgi traffic, as a clotrimazole (CTZ)-resistance gene in a multicopy library screen. Multicopies of the YPT31 homolog YPT32 also conferred resistance to CTZ, and single disruption of YPT31 or YPT32 resulted in sensitivity to CTZ. Pdr5p, an ATP-binding cassette (ABC) transporter at the plasma membrane, was the most important factor for mediating basal resistance to CTZ, suggesting that Ypt31p and Ypt32p might be involved in the trafficking of Pdr5p to the plasma membrane. However, the activity of Pdr5p was independent of YPT31 or YPT32, and multicopies of YPT31 or YPT32 still conferred resistance to CTZ in pdr5 cells. To elucidate the roles of YPT31 and YPT32 in CTZ resistance, we analyzed mutants of 11 genes that are involved in the following vesicular trafficking: Golgi traffic (kes1, trs33, trs65, gyp1, trs85, and gyp2), vacuole inheritance (ypt7), endocytosis (rcy1 and ypt51) and exocytosis (msb3 and msb4). All of the mutant cells except ypt51, msb3 and msb4 were sensitive to CTZ, indicating that vacuoles were involved in CTZ resistance, since vacuole formation requires proper Golgi-trafficking and endocytosis. Microscopic analysis showed abnormal vacuoles in ypt31 cells. Multicopies of YPT31 or YPT32 conferred resistance to CTZ in AD1-8 cells, which are defective in seven major drug transporters, and in pdr5 ypt7 cells, but not in ypt7 or AD1-8-7 (AD1-8/ypt7) cells. These results indicated that Ypt31p and Ypt32p played minor but compensatory roles in cellular resistance to CTZ through vacuoles and specific ABC transporter(s) other than Pdr5p. Copyright © 2012 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  16. Fasting Induces Nuclear Factor E2-Related Factor 2 and ATP-Binding Cassette Transporters via Protein Kinase A and Sirtuin-1 in Mouse and Human

    Science.gov (United States)

    Kulkarni, Supriya R.; Donepudi, Ajay C.; Xu, Jialin; Wei, Wei; Cheng, Qiuqiong C.; Driscoll, Maureen V.; Johnson, Delinda A.; Johnson, Jeffrey A.; Li, Xiaoling

    2014-01-01

    Abstract Aims: The purpose of this study was to determine whether 3′-5′-cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) and Sirtuin-1 (SIRT1) dependent mechanisms modulate ATP-binding Cassette (ABC) transport protein expression. ABC transport proteins (ABCC2–4) are essential for chemical elimination from hepatocytes and biliary excretion. Nuclear factor-E2 related-factor 2 (NRF2) is a transcription factor that mediates ABCC induction in response to chemical inducers and liver injury. However, a role for NRF2 in the regulation of transporter expression in nonchemical models of liver perturbation is largely undescribed. Results: Here we show that fasting increased NRF2 target gene expression through NRF2- and SIRT1–dependent mechanisms. In intact mouse liver, fasting induces NRF2 target gene expression by at least 1.5 to 5-fold. In mouse and human hepatocytes, treatment with 8-Bromoadenosine-cAMP, a cAMP analogue, increased NRF2 target gene expression and antioxidant response element activity, which was decreased by the PKA inhibitor, H-89. Moreover, fasting induced NRF2 target gene expression was decreased in liver and hepatocytes of SIRT1 liver-specific null mice and NRF2-null mice. Lastly, NRF2 and SIRT1 were recruited to MAREs and Antioxidant Response Elements (AREs) in the human ABCC2 promoter. Innovation: Oxidative stress mediated NRF2 activation is well described, yet the influence of basic metabolic processes on NRF2 activation is just emerging. Conclusion: The current data point toward a novel role of nutrient status in regulation of NRF2 activity and the antioxidant response, and indicates that cAMP/PKA and SIRT1 are upstream regulators for fasting-induced activation of the NRF2-ARE pathway. Antioxid. Redox Signal. 20, 15–30. PMID:23725046

  17. Galectin-3 silencing inhibits epirubicin-induced ATP binding cassette transporters and activates the mitochondrial apoptosis pathway via β-catenin/GSK-3β modulation in colorectal carcinoma.

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    Yung-Kuo Lee

    Full Text Available Multidrug resistance (MDR, an unfavorable factor compromising the treatment efficacy of anticancer drugs, involves the upregulation of ATP binding cassette (ABC transporters and induction of galectin-3 signaling. Galectin-3 plays an anti-apoptotic role in many cancer cells and regulates various pathways to activate MDR. Thus, the inhibition of galectin-3 has the potential to enhance the efficacy of the anticancer drug epirubicin. In this study, we examined the effects and mechanisms of silencing galectin-3 via RNA interference (RNAi on the β-catenin/GSK-3β pathway in human colon adenocarcinoma Caco-2 cells. Galectin-3 knockdown increased the intracellular accumulation of epirubicin in Caco-2 cells; suppressed the mRNA expression of galectin-3, β-catenin, cyclin D1, c-myc, P-glycoprotein (P-gp, MDR-associated protein (MRP 1, and MRP2; and downregulated the protein expression of P-gp, cyclin D1, galectin-3, β-catenin, c-Myc, and Bcl-2. Moreover, galectin-3 RNAi treatment significantly increased the mRNA level of GSK-3β, Bax, caspase-3, and caspase-9; remarkably increased the Bax-to-Bcl-2 ratio; and upregulated the GSK-3β and Bax protein expressions. Apoptosis was induced by galectin-3 RNAi and/or epirubicin as demonstrated by chromatin condensation, a higher sub-G1 phase proportion, and increased caspase-3 and caspase-9 activity, indicating an intrinsic/mitochondrial apoptosis pathway. Epirubicin-mediated resistance was effectively inhibited via galectin-3 RNAi treatment. However, these phenomena could be rescued after galectin-3 overexpression. We show for the first time that the silencing of galectin-3 sensitizes MDR cells to epirubicin by inhibiting ABC transporters and activating the mitochondrial pathway of apoptosis through modulation of the β-catenin/GSK-3β pathway in human colon cancer cells.

  18. ATP-Binding Cassette Transporter VcaM from Vibrio cholerae is Dependent on the Outer Membrane Factor Family for Its Function

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    Wen-Jung Lu

    2018-03-01

    Full Text Available Vibrio cholerae ATP-binding cassette transporter VcaM (V. cholerae ABC multidrug resistance pump has previously been shown to confer resistance to a variety of medically important drugs. In this study, we set to analyse its properties both in vitro in detergent-solubilised state and in vivo to differentiate its dependency on auxiliary proteins for its function. We report the first detailed kinetic parameters of purified VcaM and the rate of phosphate (Pi production. To determine the possible functional dependencies of VcaM on the tripartite efflux pumps we then utilized different E. coli strains lacking the principal secondary transporter AcrB (Acriflavine resistance protein, as well as cells lacking the outer membrane factor (OMF TolC (Tolerance to colicins. Consistent with the ATPase function of VcaM we found it to be susceptible to sodium orthovanadate (NaOV, however, we also found a clear dependency of VcaM function on TolC. Inhibitors targeting secondary active transporters had no effects on either VcaM-conferred resistance or Hoechst 33342 accumulation, suggesting that VcaM might be capable of engaging with the TolC-channel without periplasmic mediation by additional transporters. Our findings are indicative of VcaM being capable of a one-step substrate translocation from cytosol to extracellular space utilising the TolC-channel, making it the only multidrug ABC-transporter outside of the MacB-family with demonstrable TolC-dependency.

  19. Suppression of c-Myc is involved in multi-walled carbon nanotubes' down-regulation of ATP-binding cassette transporters in human colon adenocarcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Zhaojing [Department of Pharmacology, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, 430030 Wuhan (China); Xu, Yonghong [Institute of Ophthalmological Research, Department of Ophthalmology, Renmin Hospital of Wuhan University, 430060 Wuhan (China); Meng, Xiangning [School of Materials and Metallurgy, Northeastern University, Shenyang 110819 (China); Watari, Fumio [Department of Biomedical, Dental Materials and Engineering, Graduate School of Dental Medicine, Hokkaido University, Sapporo 060-8586 (Japan); Liu, Hudan, E-mail: hudanliu@hust.edu.cn [Department of Pharmacology, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, 430030 Wuhan (China); Chen, Xiao, E-mail: mornsmile@yahoo.com [Department of Pharmacology, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, 430030 Wuhan (China)

    2015-01-01

    Over-expression of ATP-binding cassette (ABC) transporters, a large family of integral membrane proteins that decrease cellular drug uptake and accumulation by active extrusion, is one of the major causes of cancer multi-drug resistance (MDR) that frequently leads to failure of chemotherapy. Carbon nanotubes (CNTs)-based drug delivery devices hold great promise in enhancing the efficacy of cancer chemotherapy. However, CNTs' effects on the ABC transporters remain under-investigated. In this study, we found that multiwalled carbon nanotubes (MWCNTs) reduced transport activity and expression of ABC transporters including ABCB1/Pgp and ABCC4/MRP4 in human colon adenocarcinoma Caco-2 cells. Proto-oncogene c-Myc, which directly regulates ABC gene expression, was concurrently decreased in MWCNT-treated cells and forced over-expression of c-Myc reversed MWCNTs' inhibitory effects on ABCB1 and ABCC4 expression. MWCNT-cell membrane interaction and cell membrane oxidative damage were observed. However, antioxidants such as vitamin C, β-mecaptoethanol and dimethylthiourea failed to antagonize MWCNTs' down-regulation of ABC transporters. These data suggest that MWCNTs may act on c-Myc, but not through oxidative stress, to down-regulate ABC transporter expression. Our findings thus shed light on CNTs' novel cellular effects that may be utilized to develop CNTs-based drug delivery devices to overcome ABC transporter-mediated cancer chemoresistance.

  20. Efficient purification and reconstitution of ATP binding cassette transporter B6 (ABCB6) for functional and structural studies.

    Science.gov (United States)

    Chavan, Hemantkumar; Khan, Mohiuddin Md Taimur; Tegos, George; Krishnamurthy, Partha

    2013-08-02

    The mitochondrial ATP binding cassette transporter ABCB6 has been associated with a broad range of physiological functions, including growth and development, therapy-related drug resistance, and the new blood group system Langereis. ABCB6 has been proposed to regulate heme synthesis by shuttling coproporphyrinogen III from the cytoplasm into the mitochondria. However, direct functional information of the transport complex is not known. To understand the role of ABCB6 in mitochondrial transport, we developed an in vitro system with pure and active protein. ABCB6 overexpressed in HEK293 cells was solubilized from mitochondrial membranes and purified to homogeneity. Purified ABCB6 showed a high binding affinity for MgATP (Kd = 0.18 μM) and an ATPase activity with a Km of 0.99 mM. Reconstitution of ABCB6 into liposomes allowed biochemical characterization of the ATPase including (i) substrate-stimulated ATPase activity, (ii) transport kinetics of its proposed endogenous substrate coproporphyrinogen III, and (iii) transport kinetics of substrates identified using a high throughput screening assay. Mutagenesis of the conserved lysine to alanine (K629A) in the Walker A motif abolished ATP hydrolysis and substrate transport. These results suggest a direct interaction between mitochondrial ABCB6 and its transport substrates that is critical for the activity of the transporter. Furthermore, the simple immunoaffinity purification of ABCB6 to near homogeneity and efficient reconstitution of ABCB6 into liposomes might provide the basis for future studies on the structure/function of ABCB6.

  1. A Conserved Mitochondrial ATP-binding Cassette Transporter Exports Glutathione Polysulfide for Cytosolic Metal Cofactor Assembly*♦

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    Schaedler, Theresia A.; Thornton, Jeremy D.; Kruse, Inga; Schwarzländer, Markus; Meyer, Andreas J.; van Veen, Hendrik W.; Balk, Janneke

    2014-01-01

    An ATP-binding cassette transporter located in the inner mitochondrial membrane is involved in iron-sulfur cluster and molybdenum cofactor assembly in the cytosol, but the transported substrate is unknown. ATM3 (ABCB25) from Arabidopsis thaliana and its functional orthologue Atm1 from Saccharomyces cerevisiae were expressed in Lactococcus lactis and studied in inside-out membrane vesicles and in purified form. Both proteins selectively transported glutathione disulfide (GSSG) but not reduced glutathione in agreement with a 3-fold stimulation of ATPase activity by GSSG. By contrast, Fe2+ alone or in combination with glutathione did not stimulate ATPase activity. Arabidopsis atm3 mutants were hypersensitive to an inhibitor of glutathione biosynthesis and accumulated GSSG in the mitochondria. The growth phenotype of atm3-1 was strongly enhanced by depletion of the mitochondrion-localized, GSH-dependent persulfide oxygenase ETHE1, suggesting that the physiological substrate of ATM3 contains persulfide in addition to glutathione. Consistent with this idea, a transportomics approach using mass spectrometry showed that glutathione trisulfide (GS-S-SG) was transported by Atm1. We propose that mitochondria export glutathione polysulfide, containing glutathione and persulfide, for iron-sulfur cluster assembly in the cytosol. PMID:25006243

  2. Disruption of lolCDE, Encoding an ATP-Binding Cassette Transporter, Is Lethal for Escherichia coli and Prevents Release of Lipoproteins from the Inner Membrane

    OpenAIRE

    Narita, Shin-ichiro; Tanaka, Kimie; Matsuyama, Shin-ichi; Tokuda, Hajime

    2002-01-01

    ATP-binding cassette transporter LolCDE was previously identified, by using reconstituted proteoliposomes, as an apparatus catalyzing the release of outer membrane-specific lipoproteins from the inner membrane of Escherichia coli. Mutations resulting in defective LolD were previously shown to be lethal for E. coli. The amino acid sequences of LolC and LolE are similar to each other, but the necessity of both proteins for lipoprotein release has not been proved. Moreover, previous reconstituti...

  3. Prenatal Ethanol Exposure Up-Regulates the Cholesterol Transporters ATP-Binding Cassette A1 and G1 and Reduces Cholesterol Levels in the Developing Rat Brain

    OpenAIRE

    Zhou, Chunyan; Chen, Jing; Zhang, Xiaolu; Costa, Lucio G.; Guizzetti, Marina

    2014-01-01

    Aims: Cholesterol plays a pivotal role in many aspects of brain development; reduced cholesterol levels during brain development, as a consequence of genetic defects in cholesterol biosynthesis, leads to severe brain damage, including microcephaly and mental retardation, both of which are also hallmarks of the fetal alcohol syndrome. We had previously shown that ethanol up-regulates the levels of two cholesterol transporters, ABCA1 (ATP binding cassette-A1) and ABCG1, leading to increased cho...

  4. A role for calcium in the regulation of ATP-binding cassette, sub-family C, member 3 (ABCC3) gene expression in a model of epidermal growth factor-mediated breast cancer epithelial-mesenchymal transition.

    Science.gov (United States)

    Stewart, Teneale A; Azimi, Iman; Thompson, Erik W; Roberts-Thomson, Sarah J; Monteith, Gregory R

    2015-03-13

    Epithelial-mesenchymal transition (EMT), a process implicated in cancer metastasis, is associated with the transcriptional regulation of members of the ATP-binding cassette superfamily of efflux pumps, and drug resistance in breast cancer cells. Epidermal growth factor (EGF)-induced EMT in MDA-MB-468 breast cancer cells is calcium signal dependent. In this study induction of EMT was shown to result in the transcriptional up-regulation of ATP-binding cassette, subfamily C, member 3 (ABCC3), a member of the ABC transporter superfamily, which has a recognized role in multidrug resistance. Buffering of cytosolic free calcium inhibited EGF-mediated ABCC3 increases, indicating a calcium-dependent mode of regulation. Silencing of TRPM7 (an ion channel involved in EMT associated vimentin induction) did not inhibit ABCC3 up-regulation. Silencing of the store operated calcium entry (SOCE) pathway components ORAI1 and STIM1 also did not alter ABCC3 induction by EGF. However, the calcium permeable ion channel transient receptor potential cation channel, subfamily C, member 1 (TRPC1) appears to contribute to the regulation of both basal and EGF-induced ABCC3 mRNA. Improved understanding of the relationship between calcium signaling, EMT and the regulation of genes important in therapeutic resistance may help identify novel therapeutic targets for breast cancer. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. Regulation of Human γδ T Cells by BTN3A1 Protein Stability and ATP-Binding Cassette Transporters

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    David A. Rhodes

    2018-04-01

    Full Text Available Activation of human Vγ9/Vδ2 T cells by “phosphoantigens” (pAg, the microbial metabolite (E-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMB-PP and the endogenous isoprenoid intermediate isopentenyl pyrophosphate, requires expression of butyrophilin BTN3A molecules by presenting cells. However, the precise mechanism of activation of Vγ9/Vδ2 T cells by BTN3A molecules remains elusive. It is not clear what conformation of the three BTN3A isoforms transmits activation signals nor how externally delivered pAg accesses the cytosolic B30.2 domain of BTN3A1. To approach these problems, we studied two HLA haplo-identical HeLa cell lines, termed HeLa-L and HeLa-M, which showed marked differences in pAg-dependent stimulation of Vγ9/Vδ2 T cells. Levels of IFN-γ secretion by Vγ9/Vδ2 T cells were profoundly increased by pAg loading, or by binding of the pan-BTN3A specific agonist antibody CD277 20.1, in HeLa-M compared to HeLa-L cells. IL-2 production from a murine hybridoma T cell line expressing human Vγ9/Vδ2 T cell receptor (TCR transgenes confirmed that the differential responsiveness to HeLa-L and HeLa-M was TCR dependent. By tissue typing, both HeLa lines were shown to be genetically identical and full-length transcripts of the three BTN3A isoforms were detected in equal abundance with no sequence variation. Expression of BTN3A and interacting molecules, such as periplakin or RhoB, did not account for the functional variation between HeLa-L and HeLa-M cells. Instead, the data implicate a checkpoint controlling BTN3A1 stability and protein trafficking, acting at an early time point in its maturation. In addition, plasma membrane profiling was used to identify proteins upregulated in HMB-PP-treated HeLa-M. ABCG2, a member of the ATP-binding cassette (ABC transporter family was the most significant candidate, which crucially showed reduced expression in HeLa-L. Expression of a subset of ABC transporters, including ABCA1 and ABCG1, correlated

  6. Overexpression of the ATP binding cassette gene ABCA1 determines resistance to Curcumin in M14 melanoma cells.

    Science.gov (United States)

    Bachmeier, Beatrice E; Iancu, Cristina M; Killian, Peter H; Kronski, Emanuel; Mirisola, Valentina; Angelini, Giovanna; Jochum, Marianne; Nerlich, Andreas G; Pfeffer, Ulrich

    2009-12-23

    Curcumin induces apoptosis in many cancer cells and it reduces xenograft growth and the formation of lung metastases in nude mice. Moreover, the plant derived polyphenol has been reported to be able to overcome drug resistance to classical chemotherapy. These features render the drug a promising candidate for tumor therapy especially for cancers known for their high rates concerning therapy resistance like melanoma. We show here that the melanoma cell line M14 is resistant to Curcumin induced apoptosis, which correlates with the absence of any effect on NFkappaB signaling. We show that CXCL1 a chemokine that is down regulated in breast cancer cells by Curcumin in an NFkappaB dependent manner is expressed at variable levels in human melanomas. Yet in M14 cells, CXCL1 expression did not change upon Curcumin treatment. Following the hypothesis that Curcumin is rapidly removed from the resistant cells, we analyzed expression of known multi drug resistance genes and cellular transporters in M14 melanoma cells and in the Curcumin sensitive breast cancer cell line MDA-MB-231. ATP-binding cassette transporter ABCA1, a gene involved in the cellular lipid removal pathway is over-expressed in resistant M14 melanoma as compared to the sensitive MDA-MB-231 breast cancer cells. Gene silencing of ABCA1 by siRNA sensitizes M14 cells to the apoptotic effect of Curcumin most likely as a result of reduced basal levels of active NFkappaB. Moreover, ABCA1 silencing alone also induces apoptosis and reduces p65 expression. Resistance to Curcumin thus follows classical pathways and ABCA1 expression should be considered as response marker.

  7. Overexpression of the ATP binding cassette gene ABCA1 determines resistance to Curcumin in M14 melanoma cells

    Directory of Open Access Journals (Sweden)

    Angelini Giovanna

    2009-12-01

    Full Text Available Abstract Background Curcumin induces apoptosis in many cancer cells and it reduces xenograft growth and the formation of lung metastases in nude mice. Moreover, the plant derived polyphenol has been reported to be able to overcome drug resistance to classical chemotherapy. These features render the drug a promising candidate for tumor therapy especially for cancers known for their high rates concerning therapy resistance like melanoma. Results We show here that the melanoma cell line M14 is resistant to Curcumin induced apoptosis, which correlates with the absence of any effect on NFκB signaling. We show that CXCL1 a chemokine that is down regulated in breast cancer cells by Curcumin in an NFκB dependant manner is expressed at variable levels in human melanomas. Yet in M14 cells, CXCL1 expression did not change upon Curcumin treatment. Following the hypothesis that Curcumin is rapidly removed from the resistant cells, we analyzed expression of known multi drug resistance genes and cellular transporters in M14 melanoma cells and in the Curcumin sensitive breast cancer cell line MDA-MB-231. ATP-binding cassette transporter ABCA1, a gene involved in the cellular lipid removal pathway is over-expressed in resistant M14 melanoma as compared to the sensitive MDA-MB-231 breast cancer cells. Gene silencing of ABCA1 by siRNA sensitizes M14 cells to the apoptotic effect of Curcumin most likely as a result of reduced basal levels of active NFκB. Moreover, ABCA1 silencing alone also induces apoptosis and reduces p65 expression. Conclusion Resistance to Curcumin thus follows classical pathways and ABCA1 expression should be considered as response marker.

  8. ATP binding cassette A1 (ABCA1) mediates microparticle formation during high-density lipoprotein (HDL) biogenesis.

    Science.gov (United States)

    Hafiane, Anouar; Genest, Jacques

    2017-02-01

    Micro-particles (MP) are secreted by various cells. Their biological roles in health and in disease remain unknown. Here we describe formation of MP in the process of ABCA1-dependent cholesterol efflux in different cell types. The ATP-binding cassette transporter, subfamily A, member 1 (ABCA1) is the rate-limiting step in the biogenesis of high-density lipoproteins (HDL). We have found that ABCA1 and apoA-I contribute to the formation of MP. Using cell-based systems with overexpression and selective inactivation of ABCA1, pharmacological blockade and modulation of membrane cholesterol content, we characterized MP release from various cell lines. We studied MP release in BHK cells stably expressing ABCA1 under mifepristone control, human THP-1 macrophages and HepG2 cells without, or with incubation with human apoA-I. ABCA1 mediates the production of MPs containing cholesterol. This was also confirmed in primary human monocyte-derived macrophages (MDMs). Adding apoA-I markedly increases MP release from cells. Inhibition of ABCA1 with probucol or decreasing plasma membrane cholesterol with methyl-β cyclodextrin (CDX) markedly reduced MP release and nascent HDL formation. MPs do not contain apoA-I, but contain flotilin-2, a marker of plasma membrane, and CD63, an exosome marker. MPs exhibit considerable size heterogeneity (50-250 nm). We show that MPs are lipoprotein-sized structures created by the ABCA1 transporter, and contribute approximately 30% of ABCA1-and apoA-I mediated cholesterol efflux. In addition, we found that MPs release from cells consists, in part, of exosomes and depends on the same pathway used for HDL biogenesis. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. ATP binding and hydrolysis disrupt the high-affinity interaction between the heme ABC transporter HmuUV and its cognate substrate-binding protein.

    Science.gov (United States)

    Qasem-Abdullah, Hiba; Perach, Michal; Livnat-Levanon, Nurit; Lewinson, Oded

    2017-09-01

    Using the energy of ATP hydrolysis, ABC transporters catalyze the trans-membrane transport of molecules. In bacteria, these transporters partner with a high-affinity substrate-binding protein (SBP) to import essential micronutrients. ATP binding by Type I ABC transporters (importers of amino acids, sugars, peptides, and small ions) stabilizes the interaction between the transporter and the SBP, thus allowing transfer of the substrate from the latter to the former. In Type II ABC transporters (importers of trace elements, e.g. vitamin B 12 , heme, and iron-siderophores) the role of ATP remains debatable. Here we studied the interaction between the Yersinia pestis ABC heme importer (HmuUV) and its partner substrate-binding protein (HmuT). Using real-time surface plasmon resonance experiments and interaction studies in membrane vesicles, we find that in the absence of ATP the transporter and the SBP tightly bind. Substrate in excess inhibits this interaction, and ATP binding by the transporter completely abolishes it. To release the stable docked SBP from the transporter hydrolysis of ATP is required. Based on these results we propose a mechanism for heme acquisition by HmuUV-T where the substrate-loaded SBP docks to the nucleotide-free outward-facing conformation of the transporter. ATP binding leads to formation of an occluded state with the substrate trapped in the trans-membrane translocation cavity. Subsequent ATP hydrolysis leads to substrate delivery to the cytoplasm, release of the SBP, and resetting of the system. We propose that other Type II ABC transporters likely share the fundamentals of this mechanism. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  10. Structural elucidation of transmembrane domain zero (TMD0) of EcdL: A multidrug resistance-associated protein (MRP) family of ATP-binding cassette transporter protein revealed by atomistic simulation.

    Science.gov (United States)

    Bera, Krishnendu; Rani, Priyanka; Kishor, Gaurav; Agarwal, Shikha; Kumar, Antresh; Singh, Durg Vijay

    2017-09-20

    ATP-Binding cassette (ABC) transporters play an extensive role in the translocation of diverse sets of biologically important molecules across membrane. EchnocandinB (antifungal) and EcdL protein of Aspergillus rugulosus are encoded by the same cluster of genes. Co-expression of EcdL and echinocandinB reflects tightly linked biological functions. EcdL belongs to Multidrug Resistance associated Protein (MRP) subfamily of ABC transporters with an extra transmembrane domain zero (TMD0). Complete structure of MRP subfamily comprising of TMD0 domain, at atomic resolution is not known. We hypothesized that the transportation of echonocandinB is mediated via EcdL protein. Henceforth, it is pertinent to know the topological arrangement of TMD0, with other domains of protein and its possible role in transportation of echinocandinB. Absence of effective template for TMD0 domain lead us to model by I-TASSER, further structure has been refined by multiple template modelling using homologous templates of remaining domains (TMD1, NBD1, TMD2, NBD2). The modelled structure has been validated for packing, folding and stereochemical properties. MD simulation for 0.1 μs has been carried out in the biphasic environment for refinement of modelled protein. Non-redundant structures have been excavated by clustering of MD trajectory. The structural alignment of modelled structure has shown Z-score -37.9; 31.6, 31.5 with RMSD; 2.4, 4.2, 4.8 with ABC transporters; PDB ID 4F4C, 4M1 M, 4M2T, respectively, reflecting the correctness of structure. EchinocandinB has been docked to the modelled as well as to the clustered structures, which reveals interaction of echinocandinB with TMD0 and other TM helices in the translocation path build of TMDs.

  11. Crystal structures and mutational analysis of the arginine-, lysine-, histidine-binding protein ArtJ from Geobacillus stearothermophilus. Implications for interactions of ArtJ with its cognate ATP-binding cassette transporter, Art(MP)2.

    Science.gov (United States)

    Vahedi-Faridi, Ardeschir; Eckey, Viola; Scheffel, Frank; Alings, Claudia; Landmesser, Heidi; Schneider, Erwin; Saenger, Wolfram

    2008-01-11

    ArtJ is the substrate-binding component (receptor) of the ATP-binding cassette (ABC) transport system ArtJ-(MP)(2) from the thermophilic bacterium Geobacillus stearothermophilus that is specific for arginine, lysine, and histidine. The highest affinity is found for arginine (K(d)=0.039(+/-0.014) microM), while the affinities for lysine and histidine are about tenfold lower. We have determined the X-ray structures of ArtJ liganded with each of these substrates at resolutions of 1.79 A (arginine), 1.79 A (lysine), and 2.35 A (histidine), respectively. As found for other solute receptors, the polypeptide chain is folded into two distinct domains (lobes) connected by a hinge. The interface between the lobes forms the substrate-binding pocket whose geometry is well preserved in all three ArtJ/amino acid complexes. Structure-derived mutational analyses indicated the crucial role of a region in the carboxy-terminal lobe of ArtJ in contacting the transport pore Art(MP)(2) and revealed the functional importance of Gln132 and Trp68. While variant Gln132Leu exhibited lower binding affinity for arginine but no binding of lysine and histidine, the variant Trp68Leu had lost binding activity for all three substrates. The results are discussed in comparison with known structures of homologous proteins from mesophilic bacteria.

  12. Lobular Distribution and Variability in Hepatic ATP Binding Cassette Protein B1 (ABCB1, P-gp: Ontogenetic Differences and Potential for Toxicity

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    Ngu Njei Abanda

    2017-02-01

    Full Text Available The ATP Binding Cassette B1 (ABCB1 transporter has critical roles in endo- and xenobiotic efficacy and toxicity. To understand population variability in hepatic transport we determined ABCB1 mRNA and protein levels in total liver lysates sampled from 8 pre-defined sites (n = 24, 18–69 years, and in S9 from randomly acquired samples (n = 87, 7 days–87 years. ABCB1 levels did not differ significantly throughout individual livers and showed 4.4-fold protein variation between subjects. Neither mRNA nor protein levels varied with sex, ethnicity, obesity or triglycerides in lysates or S9 (that showed the same relationships, but protein levels were lower in pediatric S9 (p < 0.0001, with 76% of adult ABCB1 present at birth and predicted to mature in 5 years. Pediatric total liver lysates were not available. In summary, opportunistic collection for studying human hepatic ABCB1 is acceptable. Additionally, ABCB1 may be lower in children, indicating differential potential for toxicity and response to therapy in this special population.

  13. ATP-binding cassette G-subfamily transporter 2 regulates cell cycle progression and asymmetric division in mouse cardiac side population progenitor cells.

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    Sereti, Konstantina-Ioanna; Oikonomopoulos, Angelos; Unno, Kazumasa; Cao, Xin; Qiu, Yiling; Liao, Ronglih

    2013-01-04

    After cardiac injury, cardiac progenitor cells are acutely reduced and are replenished in part by regulated self-renewal and proliferation, which occurs through symmetric and asymmetric cellular division. Understanding the molecular cues controlling progenitor cell self-renewal and lineage commitment is critical for harnessing these cells for therapeutic regeneration. We previously have found that the cell surface ATP-binding cassette G-subfamily transporter 2 (Abcg2) influences the proliferation of cardiac side population (CSP) progenitor cells, but through unclear mechanisms. To determine the role of Abcg2 on cell cycle progression and mode of division in mouse CSP cells. Herein, using CSP cells isolated from wild-type and Abcg2 knockout mice, we found that Abcg2 regulates G1-S cell cycle transition by fluorescence ubiquitination cell cycle indicators, cell cycle-focused gene expression arrays, and confocal live-cell fluorescent microscopy. Moreover, we found that modulation of cell cycle results in transition from symmetric to asymmetric cellular division in CSP cells lacking Abcg2. Abcg2 modulates CSP cell cycle progression and asymmetric cell division, establishing a mechanistic link between this surface transporter and cardiac progenitor cell function. Greater understanding of progenitor cell biology and, in particular, the regulation of resident progenitor cell homeostasis is vital for guiding the future development of cell-based therapies for cardiac regeneration.

  14. Crystal structure of the peptidase domain of Streptococcus ComA, a bifunctional ATP-binding cassette transporter involved in the quorum-sensing pathway.

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    Ishii, Seiji; Yano, Takato; Ebihara, Akio; Okamoto, Akihiro; Manzoku, Miho; Hayashi, Hideyuki

    2010-04-02

    ComA of Streptococcus is a member of the bacteriocin-associated ATP-binding cassette transporter family and is postulated to be responsible for both the processing of the propeptide ComC and secretion of the mature quorum-sensing signal. The 150-amino acid peptidase domain (PEP) of ComA specifically recognizes an extended region of ComC that is 15 amino acids in length. It has been proposed that an amphipathic alpha-helix formed by the N-terminal leader region of ComC, as well as the Gly-Gly motif at the cleavage site, is critical for the PEP-ComC interaction. To elucidate the substrate recognition mechanism, we determined the three-dimensional crystal structure of Streptococcus mutans PEP and then constructed models for the PEP.ComC complexes. PEP had an overall structure similar to the papain-like cysteine proteases as has long been predicted. The active site was located at the bottom of a narrow cleft, which is suitable for binding the Gly-Gly motif. Together with the results from mutational experiments, a shallow hydrophobic concave surface of PEP was proposed as a site that accommodates the N-terminal helix of ComC. This dual mode of substrate recognition would provide the small PEP domain with an extremely high substrate specificity.

  15. Lipid Absorption Defects in Intestine-specific Microsomal Triglyceride Transfer Protein and ATP-binding Cassette Transporter A1-deficient Mice*

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    Iqbal, Jahangir; Parks, John S.; Hussain, M. Mahmood

    2013-01-01

    We have previously described apolipoprotein B (apoB)-dependent and -independent cholesterol absorption pathways and the role of microsomal triglyceride transfer protein (MTP) and ATP-binding cassette transporter A1 (ABCA1) in these pathways. To assess the contribution of these pathways to cholesterol absorption and to determine whether there are other pathways, we generated mice that lack MTP and ABCA1, individually and in combination, in the intestine. Intestinal deletions of Mttp and Abca1 decreased plasma cholesterol concentrations by 45 and 24%, respectively, whereas their combined deletion reduced it by 59%. Acute cholesterol absorption was reduced by 28% in the absence of ABCA1, and it was reduced by 92–95% when MTP was deleted in the intestine alone or together with ABCA1. MTP deficiency significantly reduced triglyceride absorption, although ABCA1 deficiency had no effect. ABCA1 deficiency did not affect cellular lipids, but Mttp deficiency significantly increased intestinal levels of triglycerides and free fatty acids. Accumulation of intestinal free fatty acids, but not triglycerides, in Mttp-deficient intestines was prevented when mice were also deficient in intestinal ABCA1. Combined deficiency of these genes increased intestinal fatty acid oxidation as a consequence of increased expression of peroxisome proliferator-activated receptor-γ (PPARγ) and carnitine palmitoyltransferase 1α (CPT1α). These studies show that intestinal MTP and ABCA1 are critical for lipid absorption and are the main determinants of plasma and intestinal lipid levels. Reducing their activities might lower plasma lipid concentrations. PMID:24019513

  16. Suppression of the ATP-binding cassette transporter ABCC4 impairs neuroblastoma tumour growth and sensitises to irinotecan in vivo.

    Science.gov (United States)

    Murray, Jayne; Valli, Emanuele; Yu, Denise M T; Truong, Alan M; Gifford, Andrew J; Eden, Georgina L; Gamble, Laura D; Hanssen, Kimberley M; Flemming, Claudia L; Tan, Alvin; Tivnan, Amanda; Allan, Sophie; Saletta, Federica; Cheung, Leanna; Ruhle, Michelle; Schuetz, John D; Henderson, Michelle J; Byrne, Jennifer A; Norris, Murray D; Haber, Michelle; Fletcher, Jamie I

    2017-09-01

    The ATP-binding cassette transporter ABCC4 (multidrug resistance protein 4, MRP4) mRNA level is a strong predictor of poor clinical outcome in neuroblastoma which may relate to its export of endogenous signalling molecules and chemotherapeutic agents. We sought to determine whether ABCC4 contributes to development, growth and drug response in neuroblastoma in vivo. In neuroblastoma patients, high ABCC4 protein levels were associated with reduced overall survival. Inducible knockdown of ABCC4 strongly inhibited the growth of human neuroblastoma cells in vitro and impaired the growth of neuroblastoma xenografts. Loss of Abcc4 in the Th-MYCN transgenic neuroblastoma mouse model did not impact tumour formation; however, Abcc4-null neuroblastomas were strongly sensitised to the ABCC4 substrate drug irinotecan. Our findings demonstrate a role for ABCC4 in neuroblastoma cell proliferation and chemoresistance and provide rationale for a strategy where inhibition of ABCC4 should both attenuate the growth of neuroblastoma and sensitise tumours to ABCC4 chemotherapeutic substrates. Copyright © 2017 Elsevier Ltd. All rights reserved.

  17. Flavone Glucoside Uptake into Barley Mesophyll and Arabidopsis Cell Culture Vacuoles. Energization Occurs by H+-Antiport and ATP-Binding Cassette-Type Mechanisms1

    Science.gov (United States)

    Frangne, Nathalie; Eggmann, Thomas; Koblischke, Carsten; Weissenböck, Gottfried; Martinoia, Enrico; Klein, Markus

    2002-01-01

    In many cases, secondary plant products accumulate in the large central vacuole of plant cells. However, the mechanisms involved in the transport of secondary compounds are only poorly understood. Here, we demonstrate that the transport mechanisms for the major barley (Hordeum vulgare) flavonoid saponarin (apigenin 6-C-glucosyl-7-O-glucoside) are different in various plant species: Uptake into barley vacuoles occurs via a proton antiport and is competitively inhibited by isovitexin (apigenin 6-C-glucoside), suggesting that both flavone glucosides are recognized by the same transporter. In contrast, the transport into vacuoles from Arabidopsis, which does not synthesize flavone glucosides, displays typical characteristics of ATP-binding cassette transporters. Transport of saponarin into vacuoles of both the species is saturable with a Km of 50 to 100 μm. Furthermore, the uptake of saponarin into vacuoles from a barley mutant exhibiting a strongly reduced flavone glucoside biosynthesis is drastically decreased when compared with the parent variety. Thus, the barley vacuolar flavone glucoside/H+ antiporter could be modulated by the availability of the substrate. We propose that different vacuolar transporters may be responsible for the sequestration of species-specific/endogenous and nonspecific/xenobiotic secondary compounds in planta. PMID:11842175

  18. Arabidopsis PEN3/PDR8, an ATP binding cassette transporter, contributes to nonhost resistance to inappropriate pathogens that enter by direct penetration.

    Science.gov (United States)

    Stein, Mónica; Dittgen, Jan; Sánchez-Rodríguez, Clara; Hou, Bi-Huei; Molina, Antonio; Schulze-Lefert, Paul; Lipka, Volker; Somerville, Shauna

    2006-03-01

    Arabidopsis thaliana is a host to the powdery mildew Erysiphe cichoracearum and nonhost to Blumeria graminis f. sp hordei, the powdery mildew pathogenic on barley (Hordeum vulgare). Screening for Arabidopsis mutants deficient in resistance to barley powdery mildew identified PENETRATION3 (PEN3). pen3 plants permitted both increased invasion into epidermal cells and initiation of hyphae by B. g. hordei, suggesting that PEN3 contributes to defenses at the cell wall and intracellularly. pen3 mutants were compromised in resistance to the necrotroph Plectosphaerella cucumerina and to two additional inappropriate biotrophs, pea powdery mildew (Erysiphe pisi) and potato late blight (Phytophthora infestans). Unexpectedly, pen3 mutants were resistant to E. cichoracearum. This resistance was salicylic acid-dependent and correlated with chlorotic patches. Consistent with this observation, salicylic acid pathway genes were hyperinduced in pen3 relative to the wild type. The phenotypes conferred by pen3 result from the loss of function of PLEIOTROPIC DRUG RESISTANCE8 (PDR8), a highly expressed putative ATP binding cassette transporter. PEN3/PDR8 tagged with green fluorescent protein localized to the plasma membrane in uninfected cells. In infected leaves, the protein concentrated at infection sites. PEN3/PDR8 may be involved in exporting toxic materials to attempted invasion sites, and intracellular accumulation of these toxins in pen3 may secondarily activate the salicylic acid pathway.

  19. Arabidopsis PEN3/PDR8, an ATP Binding Cassette Transporter, Contributes to Nonhost Resistance to Inappropriate Pathogens That Enter by Direct Penetration[W][OA

    Science.gov (United States)

    Stein, Mónica; Dittgen, Jan; Sánchez-Rodríguez, Clara; Hou, Bi-Huei; Molina, Antonio; Schulze-Lefert, Paul; Lipka, Volker; Somerville, Shauna

    2006-01-01

    Arabidopsis thaliana is a host to the powdery mildew Erysiphe cichoracearum and nonhost to Blumeria graminis f. sp hordei, the powdery mildew pathogenic on barley (Hordeum vulgare). Screening for Arabidopsis mutants deficient in resistance to barley powdery mildew identified PENETRATION3 (PEN3). pen3 plants permitted both increased invasion into epidermal cells and initiation of hyphae by B. g. hordei, suggesting that PEN3 contributes to defenses at the cell wall and intracellularly. pen3 mutants were compromised in resistance to the necrotroph Plectosphaerella cucumerina and to two additional inappropriate biotrophs, pea powdery mildew (Erysiphe pisi) and potato late blight (Phytophthora infestans). Unexpectedly, pen3 mutants were resistant to E. cichoracearum. This resistance was salicylic acid–dependent and correlated with chlorotic patches. Consistent with this observation, salicylic acid pathway genes were hyperinduced in pen3 relative to the wild type. The phenotypes conferred by pen3 result from the loss of function of PLEIOTROPIC DRUG RESISTANCE8 (PDR8), a highly expressed putative ATP binding cassette transporter. PEN3/PDR8 tagged with green fluorescent protein localized to the plasma membrane in uninfected cells. In infected leaves, the protein concentrated at infection sites. PEN3/PDR8 may be involved in exporting toxic materials to attempted invasion sites, and intracellular accumulation of these toxins in pen3 may secondarily activate the salicylic acid pathway. PMID:16473969

  20. IMB2026791, a Xanthone, Stimulates Cholesterol Efflux by Increasing the Binding of Apolipoprotein A-I to ATP-Binding Cassette Transporter A1

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    Zijian Xie

    2012-03-01

    Full Text Available It is known that the ATP-binding cassette transporter A1 (ABCA1 plays a major role in cholesterol homeostasis and high density lipoprotein (HDL metabolism. Several laboratories have demonstrated that ABCA1 binding to lipid-poor apolipoprotein A-I (apoA-I will mediate the assembly of nascent HDL and cellular cholesterol efflux, which suggests a possible receptor-ligand interaction between ABCA1 and apoA-I. In this study, a cell-based-ELISA-like high-throughput screening (HTS method was developed to identify the synthetic and natural compounds that can regulate binding activity of ABCA1 to apoA-I. The cell-based-ELISA-like high-throughput screen was conducted in a 96-well format using Chinese hamster ovary (CHO cells stably transfected with ABCA1 pIRE2-EGFP (Enhanced Green Fluorecence Protein expression vector and the known ABCA1 inhibitor glibenclamide as the antagonist control. From 2,600 compounds, a xanthone compound (IMB 2026791 was selected using this HTS assay, and it was proved as an apoA-I binding agonist to ABCA1 by a flow cytometry assay and western blot analysis. The [3H] cholesterol efflux assay of IMB2026791 treated ABCA1-CHO cells and PMA induced THP-1 macrophages (human acute monocytic leukemia cell further confirmed the compound as an accelerator of cholesterol efflux in a dose-dependent manner with an EC50 of 25.23 μM.

  1. Cooperative transcriptional activation of ATP-binding cassette sterol transporters ABCG5 and ABCG8 genes by nuclear receptors including Liver-X-Receptor

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    Su Sun Back

    2013-06-01

    Full Text Available The ATP-binding cassette transporters ABCG5 and ABCG8 formheterodimers that limit absorption of dietary sterols in theintestine and promote cholesterol elimination from the bodythrough hepatobiliary secretion. To identify cis-regulatoryelements of the two genes, we have cloned and analyzedtwenty-three evolutionary conserved region (ECR fragmentsusing the CMV-luciferase reporter system in HepG2 cells. TwoECRs were found to be responsive to the Liver-X-Receptor (LXR.Through elaborate deletion studies, regions containing putativeLXREs were identified and the binding of LXRα wasdemonstrated by EMSA and ChIP assay. When the LXREs wereinserted upstream of the intergenic promoter, synergisticactivation by LXRα/RXRα in combination with GATA4, HNF4α,and LRH-1, which had been shown to bind to the intergenicregion, was observed. In conclusion, we have identified twoLXREs in ABCG5/ABCG8 genes for the first time and proposethat these LXREs, especially in the ECR20, play major roles inregulating these genes. [BMB Reports 2013; 46(6: 322-327

  2. α-Lipoic acid ameliorates foam cell formation via liver X receptor α-dependent upregulation of ATP-binding cassette transporters A1 and G1.

    Science.gov (United States)

    Cheng, Li-Ching; Su, Kuo-Hui; Kou, Yu Ru; Shyue, Song-Kun; Ching, Li-Chieh; Yu, Yuan-Bin; Wu, Yuh-Lin; Pan, Ching-Chian; Lee, Tzong-Shyuan

    2011-01-01

    α-Lipoic acid (α-LA), a key cofactor in cellular energy metabolism, has protective activities in atherosclerosis, yet the detailed mechanisms are not fully understood. In this study, we examined whether α-LA affects foam cell formation and its underlying molecular mechanisms in murine macrophages. Treatment with α-LA markedly attenuated oxidized low-density lipoprotein (oxLDL)-mediated cholesterol accumulation in macrophages, which was due to increased cholesterol efflux. Additionally, α-LA treatment dose-dependently increased protein levels of ATP-binding cassette transporter A1 (ABCA1) and ABCG1 but had no effect on the protein expression of SR-A, CD36, or SR-BI involved in cholesterol homeostasis. Furthermore, α-LA increased the mRNA expression of ABCA1 and ABCG1. The upregulation of ABCA1 and ABCG1 by α-LA depended on liver X receptor α (LXRα), as evidenced by an increase in the nuclear levels of LXRα and LXRE-mediated luciferase activity and its prevention of the expression of ABCA1 and ABCG1 after inhibition of LXRα activity by the pharmacological inhibitor geranylgeranyl pyrophosphate (GGPP) or knockdown of LXRα expression with small interfering RNA (siRNA). Consistently, α-LA-mediated suppression of oxLDL-induced lipid accumulation was abolished by GGPP or LXRα siRNA treatment. In conclusion, LXRα-dependent upregulation of ABCA1 and ABCG1 may mediate the beneficial effect of α-LA on foam cell formation. Copyright © 2010 Elsevier Inc. All rights reserved.

  3. ATP-binding cassette transporter ABCA4 and chemical isomerization protect photoreceptor cells from the toxic accumulation of excess 11-cis-retinal

    Science.gov (United States)

    Quazi, Faraz; Molday, Robert S.

    2014-01-01

    The visual cycle is a series of enzyme-catalyzed reactions which converts all-trans-retinal to 11-cis-retinal for the regeneration of visual pigments in rod and cone photoreceptor cells. Although essential for vision, 11-cis-retinal like all-trans-retinal is highly toxic due to its highly reactive aldehyde group and has to be detoxified by either reduction to retinol or sequestration within retinal-binding proteins. Previous studies have focused on the role of the ATP-binding cassette transporter ABCA4 associated with Stargardt macular degeneration and retinol dehydrogenases (RDH) in the clearance of all-trans-retinal from photoreceptors following photoexcitation. How rod and cone cells prevent the accumulation of 11-cis-retinal in photoreceptor disk membranes in excess of what is required for visual pigment regeneration is not known. Here we show that ABCA4 can transport N-11-cis-retinylidene-phosphatidylethanolamine (PE), the Schiff-base conjugate of 11-cis-retinal and PE, from the lumen to the cytoplasmic leaflet of disk membranes. This transport function together with chemical isomerization to its all-trans isomer and reduction to all-trans-retinol by RDH can prevent the accumulation of excess 11-cis-retinal and its Schiff-base conjugate and the formation of toxic bisretinoid compounds as found in ABCA4-deficient mice and individuals with Stargardt macular degeneration. This segment of the visual cycle in which excess 11-cis-retinal is converted to all-trans-retinol provides a rationale for the unusually high content of PE and its long-chain unsaturated docosahexaenoyl group in photoreceptor membranes and adds insight into the molecular mechanisms responsible for Stargardt macular degeneration. PMID:24707049

  4. Mutant Allele-Specific Uncoupling of PENETRATION3 Functions Reveals Engagement of the ATP-Binding Cassette Transporter in Distinct Tryptophan Metabolic Pathways1[OPEN

    Science.gov (United States)

    Lu, Xunli; Dittgen, Jan; Piślewska-Bednarek, Mariola; Molina, Antonio; Schneider, Bernd; Doubský, Jan; Schneeberger, Korbinian; Schulze-Lefert, Paul

    2015-01-01

    Arabidopsis (Arabidopsis thaliana) PENETRATION (PEN) genes quantitatively contribute to the execution of different forms of plant immunity upon challenge with diverse leaf pathogens. PEN3 encodes a plasma membrane-resident pleiotropic drug resistance-type ATP-binding cassette transporter and is thought to act in a pathogen-inducible and PEN2 myrosinase-dependent metabolic pathway in extracellular defense. This metabolic pathway directs the intracellular biosynthesis and activation of tryptophan-derived indole glucosinolates for subsequent PEN3-mediated efflux across the plasma membrane at pathogen contact sites. However, PEN3 also functions in abiotic stress responses to cadmium and indole-3-butyric acid (IBA)-mediated auxin homeostasis in roots, raising the possibility that PEN3 exports multiple functionally unrelated substrates. Here, we describe the isolation of a pen3 allele, designated pen3-5, that encodes a dysfunctional protein that accumulates in planta like wild-type PEN3. The specific mutation in pen3-5 uncouples PEN3 functions in IBA-stimulated root growth modulation, callose deposition induced with a conserved peptide epitope of bacterial flagellin (flg22), and pathogen-inducible salicylic acid accumulation from PEN3 activity in extracellular defense, indicating the engagement of multiple PEN3 substrates in different PEN3-dependent biological processes. We identified 4-O-β-d-glucosyl-indol-3-yl formamide (4OGlcI3F) as a pathogen-inducible, tryptophan-derived compound that overaccumulates in pen3 leaf tissue and has biosynthesis that is dependent on an intact PEN2 metabolic pathway. We propose that a precursor of 4OGlcI3F is the PEN3 substrate in extracellular pathogen defense. These precursors, the shared indole core present in IBA and 4OGlcI3F, and allele-specific uncoupling of a subset of PEN3 functions suggest that PEN3 transports distinct indole-type metabolites in distinct biological processes. PMID:26023163

  5. ATP-Binding Cassette Transporter G2 Activity in the Bovine Spermatozoa Is Modulated Along the Epididymal Duct and at Ejaculation1

    Science.gov (United States)

    Caballero, Julieta; Frenette, Gilles; D'Amours, Olivier; Dufour, Maurice; Oko, Richard; Sullivan, Robert

    2012-01-01

    During their epididymal maturation, stabilizing factors such as cholesterol sulfate are associated with the sperm plasma membrane. Cholesterol is sulfated in epididymal spermatozoa by the enzyme estrogen sulfotransferase. Because of its role in the efflux of sulfate conjugates formed intracellularly by sulfotransferases, the ATP-binding cassette membrane transporter G2 (ABCG2) might have a role in the translocation of this compound across the plasma membrane. In the present study we showed that ABCG2 is present in the plasma membrane overlaying the acrosomal region of spermatozoa recovered from testis, epididymis, and after ejaculation. Although ABCG2 is also present in epididymosomes, the transporter is not transferred to spermatozoa via this mechanism. Furthermore, although epididymal sperm ABCG2 was shown to be functional, as determined by its ability to extrude Hoechst 33342 in the presence of the specific inhibitor Fumitremorgin C, ABCG2 present in ejaculated sperm was found to be nonfunctional. Additional experiments demonstrated that phosphorylation of ABCG2 tyrosyl residues, but not its localization in lipid rafts, is the mechanism responsible for its functionality. Dephosphorylation of ABCG2 in ejaculated spermatozoa is proposed to cause a partial protein relocalization to other intracellular compartments. Prostasomes are proposed to have a role in this process because incubation with this fraction of seminal plasma induces a decrease in the amount of ABCG2 in the associated sperm membrane fraction. These results demonstrate that ABCG2 plays a role in epididymal sperm maturation, but not after ejaculation. The loss of ABCG2 function after ejaculation is proposed to be regulated by prostasomes. PMID:22441796

  6. MicroRNA-20a/b regulates cholesterol efflux through post-transcriptional repression of ATP-binding cassette transporter A1.

    Science.gov (United States)

    Liang, Bin; Wang, Xin; Song, Xiaosu; Bai, Rui; Yang, Huiyu; Yang, Zhiming; Xiao, Chuanshi; Bian, Yunfei

    2017-09-01

    ATP-binding cassette transporter A1 (ABCA1) plays a crucial role in reverse cholesterol transport and exhibits anti-atherosclerosis effects. Some microRNAs (miRs) regulate ABCA1 expression, and recent studies have shown that miR-20a/b might play a critical role in atherosclerotic diseases. Here, we attempted to clarify the potential contribution of miR-20a/b in post-transcriptional regulation of ABCA1, cholesterol efflux, and atherosclerosis. We performed bioinformatics analysis and found that miR-20a/b was highly conserved and directly bound to ABCA1 mRNA with low binding free energy. Luciferase-reporter assay also confirmed that miR-20a/b significantly reduced luciferase activity associated with the ABCA1 3' untranslated region reporter construct. Additionally, miR-20a/b decreased ABCA1 expression, which, in turn, decreased cholesterol efflux and increased cholesterol content in THP-1 and RAW 264.7 macrophage-derived foam cells. In contrast, miR-20a/b inhibitors increased ABCA1 expression and cholesterol efflux, decreased cholesterol content, and inhibited foam-cell formation. Consistent with our in vitro results, miR-20a/b-treated ApoE -/- mice showed decreased ABCA1expression in the liver and reductions of reverse cholesterol transport in vivo. Furthermore, miR-20a/b regulated the formation of nascent high-density lipoprotein and promoted atherosclerotic development, whereas miR-20a/b knockdown attenuated atherosclerotic formation. miR-20 is a new miRNA capable of targeting ABCA1 and regulating ABCA1 expression. Therefore, miR-20 inhibition constitutes a new strategy for ABCA1-based treatment of atherosclerosis. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Association Study of the ATP - Binding Cassette Transporter A1 (ABCA1 Rs2230806 Genetic Variation with Lipid Profile and Coronary Artery Disease Risk in an Iranian Population

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    Habib Ghaznavi

    2018-02-01

    Full Text Available BACKGROUND: ATP - binding cassette transporter A1 (ABCA1 plays essential roles in the biogenesis of high -density lipoprotein - cholesterol. Variations in the ABCA1 gene may influence the risk of coronary artery disease (CAD. AIM: Present study aimed to investigate the association of rs2230806 (R219K polymorphism of ABCA1 gene with the development and severity of CAD in an Iranian population. MATERIALS AND METHODS: Our study population consisted of 100 patients with angiographically confirmed CAD and 100 controls. The genotyping of R219K mutation of ABCA1 gene was determined by PCR - RFLP method. Lipid profile was determined using routine colourimetric assays. Statistical analysis was done by SPSS - 16. RESULTS: The genotypic (P = 0.024 and allelic (P = 0.001 distribution of the ABCA1 R219K polymorphism were significantly different between the two groups. In a univariate analysis (with genotype RR as the reference, the RK genotype (OR = 0.46, 95%CI = 0.25-0.86, P = 0.020 and KK genotype (OR = 0.27, 95%CI = 0.11 – 0.66, P = 0.005 was significantly associated with a decreased risk of CAD. A multiple logistic regression analysis revealed that smoking (0.008, diabetes (P = 0.023, triglyceride (P = 0.001, HDL - cholesterol (P = 0.002 and ABCA1 KK genotype (P = 0.009 were significantly and independently associated with the risk of CAD. The association between different genotypes of R219K polymorphism with lipid profile was not significant in both groups (P > 0.05. The R219K polymorphism was significantly associated with severity of CAD (P < 0.05. CONCLUSION: The carriage of K allele of ABCA1 R219K polymorphism has a protective effect on CAD risk and correlates with a decreased severity of CAD. This protective effect seems to be mediated independently of plasma lipid levels.

  8. Prenatal Ethanol Exposure Up-Regulates the Cholesterol Transporters ATP-Binding Cassette A1 and G1 and Reduces Cholesterol Levels in the Developing Rat Brain.

    Science.gov (United States)

    Zhou, Chunyan; Chen, Jing; Zhang, Xiaolu; Costa, Lucio G; Guizzetti, Marina

    2014-11-01

    Cholesterol plays a pivotal role in many aspects of brain development; reduced cholesterol levels during brain development, as a consequence of genetic defects in cholesterol biosynthesis, leads to severe brain damage, including microcephaly and mental retardation, both of which are also hallmarks of the fetal alcohol syndrome. We had previously shown that ethanol up-regulates the levels of two cholesterol transporters, ABCA1 (ATP binding cassette-A1) and ABCG1, leading to increased cholesterol efflux and decreased cholesterol content in astrocytes in vitro. In the present study we investigated whether similar effects could be seen in vivo. Pregnant Sprague-Dawley rats were fed liquid diets containing 36% of the calories from ethanol from gestational day (GD) 6 to GD 21. A pair-fed control groups and an ad libitum control group were included in the study. ABCA1 and ABCG1 protein expression and cholesterol and phospholipid levels were measured in the neocortex of female and male fetuses at GD 21. Body weights were decreased in female fetuses as a consequence of ethanol treatments. ABCA1 and ABCG1 protein levels were increased, and cholesterol levels were decreased, in the neocortex of ethanol-exposed female, but not male, fetuses. Levels of phospholipids were unchanged. Control female fetuses fed ad libitum displayed an up-regulation of ABCA1 and a decrease in cholesterol content compared with pair-fed controls, suggesting that a compensatory up-regulation of cholesterol levels may occur during food restriction. Maternal ethanol consumption may affect fetal brain development by increasing cholesterol transporters' expression and reducing brain cholesterol levels. © The Author 2014. Medical Council on Alcohol and Oxford University Press. All rights reserved.

  9. Cholesterol transporter ATP-binding cassette A1 (ABCA1) is elevated in prion disease and affects PrPC and PrPSc concentrations in cultured cells.

    Science.gov (United States)

    Kumar, Rajeev; McClain, Denise; Young, Rebecca; Carlson, George A

    2008-06-01

    Prion diseases are transmissible neurodegenerative disorders of prion protein (PrP) conformation. Prion replication by conversion of benign PrPC isoforms into disease-specific PrPSc isoforms is intimately involved in prion disease pathogenesis and may be initiated in cholesterol-rich caveolae-like domains (CLD). Concentrations of the cholesterol transporter ATP-binding cassette A1 protein (ABCA1) are elevated in pre-clinical scrapie prion-infected mice and in prion-infected cells in vitro. Elevation of ABCA1 in prion-infected brain is not a direct consequence of local PrPSc accumulation, indeed levels of ABCA1 are comparable in brain regions that differ dramatically in the amount of PrPSc. Similarly, ABCA1 concentrations are identical in normal mice, transgenic mice overexpressing PrP and PrP knockout mice. In contrast, PrPC and PrPSc levels, but not Prnp mRNA, were increased by overexpression of ABCA1 in N2a neuroblastoma cells and scrapie prion-infected N2a cells (ScN2a). Conversely, RNAi-mediated knock down of Abca1 expression decreased the concentrations of PrPC in N2a cells and of PrPSc in ScN2a cells. These results suggest that ABCA1's effects on PrPC levels are post-translational and may reflect an increase in of PrPC stability, mediated either indirectly by increasing membrane cholesterol and CLD formation or by other functions of ABCA1. The increased supply of PrPC available for conversion would lead to increased PrPSc formation.

  10. Selective ATP-Binding Cassette Subfamily C Gene Expression and Proinflammatory Mediators Released by BEAS-2B after PM2.5, Budesonide, and Cotreated Exposures

    Directory of Open Access Journals (Sweden)

    Jarline Encarnación-Medina

    2017-01-01

    Full Text Available ATP-binding cassette subfamily C (ABCC genes code for phase III metabolism proteins that translocate xenobiotic (e.g., particulate matter 2.5 (PM2.5 and drug metabolites outside the cells. IL-6 secretion is related with the activation of the ABCC transporters. This study assesses ABCC1–4 gene expression changes and proinflammatory cytokine (IL-6, IL-8 release in human bronchial epithelial cells (BEAS-2B exposed to PM2.5 organic extract, budesonide (BUD, used to control inflammation in asthmatic patients, and a cotreatment (Co-T: PM2.5 and BUD. A real-time PCR assay shows that ABCC1 was upregulated in BEAS-2B exposed after 6 and 7 hr to PM2.5 extract or BUD but downregulated after 6 hr of the Co-T. ABCC3 was downregulated after 6 hr of BUD and upregulated after 6 hr of the Co-T exposures. ABCC4 was upregulated after 5 hr of PM2.5 extract, BUD, and the Co-T exposures. The cytokine assay revealed an increase in IL-6 release by BEAS-2B exposed after 5 hr to PM2.5 extract, BUD, and the Co-T. At 7 hr, the Co-T decreases IL-6 release and IL-8 at 6 hr. In conclusion, the cotreatment showed an opposite effect on exposed BEAS-2B as compared with BUD. The results suggest an interference of the BUD therapeutic potential by PM2.5.

  11. Diosgenin inhibits atherosclerosis via suppressing the MiR-19b-induced downregulation of ATP-binding cassette transporter A1.

    Science.gov (United States)

    Lv, Yun-cheng; Yang, Jing; Yao, Feng; Xie, Wei; Tang, Yan-yan; Ouyang, Xin-ping; He, Ping-ping; Tan, Yu-lin; Li, Liang; Zhang, Min; Liu, Dan; Cayabyab, Francisco S; Zheng, Xi-Long; Tang, Chao-ke

    2015-05-01

    Diosgenin (Dgn), a structural analogue of cholesterol, has been reported to have the hypolipidemic and antiatherogenic properties, but the underlying mechanisms are not fully understood. Given the key roles of macrophages in cholesterol metabolism and atherogenesis, it is critical to investigate macrophage cholesterol efflux and development of atherosclerotic lesion after Dgn treatment. This study was designed to evaluate the potential effects of Dgn on macrophage cholesterol metabolism and the development of aortic atherosclerosis, and to explore its underlying mechanisms. Dgn significantly up-regulated the expression of ATP-binding cassette transporter A1 (ABCA1) protein, but didn't affect liver X receptor α levels in foam cells derived from human THP-1 macrophages and mouse peritoneal macrophages (MPMs) as determined by western blotting. The miR-19b levels were markedly down-regulated in Dgn-treated THP-1 macrophages/MPM-derived foam cells. Cholesterol transport assays revealed that treatment with Dgn alone or together with miR-19b inhibitor notably enhanced ABCA1-dependent cholesterol efflux, resulting in the reduced levels of total cholesterol, free cholesterol and cholesterol ester as determined by high-performance liquid chromatography. The fecal 3H-sterol originating from cholesterol-laden MPMs was increased in apolipoprotein E knockout mice treated with Dgn or both Dgn and antagomiR-19b. Treatment with Dgn alone or together with antagomiR-19b elevated plasma high-density lipoprotein levels, but reduced plasma low-density lipoprotein levels. Accordingly, aortic lipid deposition and plaque area were reduced, and collagen content and ABCA1 expression were increased in mice treated with Dgn alone or together with antagomiR-19b. However, miR-19b overexpression abrogated the lipid-lowering and atheroprotective effects induced by Dgn. The present study demonstrates that Dgn enhances ABCA1-dependent cholesterol efflux and inhibits aortic atherosclerosis

  12. MicroRNA-19b promotes macrophage cholesterol accumulation and aortic atherosclerosis by targeting ATP-binding cassette transporter A1.

    Science.gov (United States)

    Lv, Yun-Cheng; Tang, Yan-Yan; Peng, Juan; Zhao, Guo-Jun; Yang, Jing; Yao, Feng; Ouyang, Xin-Ping; He, Ping-Ping; Xie, Wei; Tan, Yu-Lin; Zhang, Min; Liu, Dan; Tang, Deng-Pei; Cayabyab, Francisco S; Zheng, Xi-Long; Zhang, Da-Wei; Tian, Guo-Ping; Tang, Chao-Ke

    2014-09-01

    Macrophage accumulation of cholesterol leads to foam cell formation which is a major pathological event of atherosclerosis. Recent studies have shown that microRNA (miR)-19b might play an important role in cholesterol metabolism and atherosclerotic diseases. Here, we have identified miR-19b binding to the 3'UTR of ATP-binding cassette transporter A1 (ABCA1) transporters, and further determined the potential roles of this novel interaction in atherogenesis. To investigate the molecular mechanisms involved in a miR-19b promotion of macrophage cholesterol accumulation and the development of aortic atherosclerosis. We performed bioinformatics analysis using online websites, and found that miR-19b was highly conserved during evolution and directly bound to ABCA1 mRNA with very low binding free energy. Luciferase reporter assay confirmed that miR-19b bound to 3110-3116 sites within ABCA1 3'UTR. MiR-19b directly regulated the expression levels of endogenous ABCA1 in foam cells derived from human THP-1 macrophages and mouse peritoneal macrophages (MPMs) as determined by qRT-PCR and western blot. Cholesterol transport assays revealed that miR-19b dramatically suppressed apolipoprotein AI-mediated ABCA1-dependent cholesterol efflux, resulting in the increased levels of total cholesterol (TC), free cholesterol (FC) and cholesterol ester (CE) as revealed by HPLC. The excretion of (3)H-cholesterol originating from cholesterol-laden MPMs into feces was decreased in mice overexpressing miR-19b. Finally, we evaluated the proatherosclerotic role of miR-19b in apolipoprotein E deficient (apoE(-/-)) mice. Treatment with miR-19b precursor reduced plasma high-density lipoprotein (HDL) levels, but increased plasma low-density lipoprotein (LDL) levels. Consistently, miR-19b precursor treatment increased aortic plaque size and lipid content, but reduced collagen content and ABCA1 expression. In contrast, treatment with the inhibitory miR-19b antisense oligonucleotides (ASO) prevented or

  13. Role of ATP binding and hydrolysis in the gating of the cystic fibrosis transmembrane conductance regulator

    Directory of Open Access Journals (Sweden)

    Taras Gout

    2012-01-01

    Full Text Available The CFTR gene is unique within the ATP-binding cassette (ABC protein family, predominantly of transporters, by coding a chloride channel. The gating mechanism of ABC proteins has been characterized by the ATP Switch model in terms cycles of dimer formation and dissociation linked to ATP binding and hydrolysis, respectively. It would be of interest to assess the extent that Cystic Fibrosis Transmembrane Conductance Regulator (CFTR, a functional channel, fits the ATP Switch model for ABC transporters. Additional transporter mechanisms, namely those of Pgp and HlyB, are discussed for perspective. Literature search of databases selected key references in comparing and contrasting the gating mechanism. CFTR is a functional chloride channel facilitating transmembrane anion flow down electrochemical gradients. A dysfunctional CFTR protein results in cystic fibrosis, a fatal pleiotropic disease currently managed symptomatically. Understanding the gating mechanism will help target drug development aimed at alleviating and curing the disease.

  14. ATP Binding and Hydrolysis Properties of ABCB10 and Their Regulation by Glutathione

    Science.gov (United States)

    Qiu, Wei; Liesa, Marc; Carpenter, Elizabeth P.; Shirihai, Orian S.

    2015-01-01

    ABCB10 (ATP binding cassette sub-family B10) is a mitochondrial inner-membrane ABC transporter. ABCB10 has been shown to protect the heart from the impact of ROS during ischemia-reperfusion and to allow for proper hemoglobin synthesis during erythroid development. ABC transporters are proteins that increase ATP binding and hydrolysis activity in the presence of the transported substrate. However, molecular entities transported by ABCB10 and its regulatory mechanisms are currently unknown. Here we characterized ATP binding and hydrolysis properties of ABCB10 by using the 8-azido-ATP photolabeling technique. This technique can identify potential ABCB10 regulators, transported substrates and amino-acidic residues required for ATP binding and hydrolysis. We confirmed that Gly497 and Lys498 in the Walker A motif, Glu624 in the Walker B motif and Gly602 in the C-Loop motif of ABCB10 are required for proper ATP binding and hydrolysis activity, as their mutation changed ABCB10 8-Azido-ATP photo-labeling. In addition, we show that the potential ABCB10 transported entity and heme precursor delta-aminolevulinic acid (dALA) does not alter 8-azido-ATP photo-labeling. In contrast, oxidized glutathione (GSSG) stimulates ATP hydrolysis without affecting ATP binding, whereas reduced glutathione (GSH) inhibits ATP binding and hydrolysis. Indeed, we detectABCB10 glutathionylation in Cys547 and show that it is one of the exposed cysteine residues within ABCB10 structure. In all, we characterize essential residues for ABCB10 ATPase activity and we provide evidence that supports the exclusion of dALA as a potential substrate directly transported by ABCB10. Last, we show the first molecular mechanism by which mitochondrial oxidative status, through GSH/GSSG, can regulate ABCB10. PMID:26053025

  15. A burst of ABC genes in the genome of the polyphagous spider mite Tetranychus urticae

    NARCIS (Netherlands)

    Dermauw, W.; Osborne, E.J.; Clark, R.M.; Grbić, M.; Tirry, L.; Van Leeuwen, T.

    2013-01-01

    Background: The ABC (ATP-binding cassette) gene superfamily is widespread across all living species. The majority of ABC genes encode ABC transporters, which are membrane-spanning proteins capable of transferring substrates across biological membranes by hydrolyzing ATP. Although ABC transporters

  16. Rosuvastatin activates ATP-binding cassette transporter A1-dependent efflux ex vivo and promotes reverse cholesterol transport in macrophage cells in mice fed a high-fat diet.

    Science.gov (United States)

    Shimizu, Tomohiko; Miura, Shin-ichiro; Tanigawa, Hiroyuki; Kuwano, Takashi; Zhang, Bo; Uehara, Yoshinari; Saku, Keijiro

    2014-10-01

    It is controversial whether statins improve high-density lipoprotein (HDL) function, which plays an important role in reverse cholesterol transport in vivo. The aim of the present study was to clarify the effects of rosuvastatin and atorvastatin on reverse cholesterol transport in macrophage cells in vivo and their underlying mechanisms. Male C57BL mice were divided into 3 groups (rosuvastatin, atorvastatin, and control groups) and orally administered rosuvastatin, atorvastatin, or placebo for 6 weeks under feeding with a 0.5% cholesterol+10% coconut oil diet. After administration, although there were no changes in plasma HDL cholesterol levels among the groups, plasma from the rosuvastatin group showed an increased ability to promote ATP-binding cassette transporter A1-mediated cholesterol efflux ex vivo. In addition, capillary electrophoresis revealed a shift in HDL toward the pre-β HDL fraction only in the rosuvastatin group. Mice in all 3 groups were intraperitoneally injected with (3)H-cholesterol-labeled and cholesterol-loaded macrophages and then were monitored for the appearance of (3)H-tracer in plasma and feces. The amount of (3)H-tracer excreted into feces during 48 hours in the rosuvastatin group was greater than that in the control group. Finally, (3)H-cholesteryl oleate-HDL was intravenously injected into all groups, blood samples were taken, and the count of (3)H-cholesterol was analyzed. Plasma (3)H-cholesteryl oleate-HDL changed similarly, and no differences in fractional catabolic rates were observed. Rosuvastatin enhanced the ATP-binding cassette transporter A1-dependent HDL efflux function of reverse cholesterol transport, and this finding highlights the potential of rosuvastatin for the regression of atherosclerosis. © 2014 American Heart Association, Inc.

  17. Genome-wide identification and evolution of ATP-binding cassette transporters in the ciliate Tetrahymena thermophila: A case of functional divergence in a multigene family

    Directory of Open Access Journals (Sweden)

    Yuan Dongxia

    2010-10-01

    Full Text Available Abstract Background In eukaryotes, ABC transporters that utilize the energy of ATP hydrolysis to expel cellular substrates into the environment are responsible for most of the efflux from cells. Many members of the superfamily of ABC transporters have been linked with resistance to multiple drugs or toxins. Owing to their medical and toxicological importance, members of the ABC superfamily have been studied in several model organisms and warrant examination in newly sequenced genomes. Results A total of 165 ABC transporter genes, constituting a highly expanded superfamily relative to its size in other eukaryotes, were identified in the macronuclear genome of the ciliate Tetrahymena thermophila. Based on ortholog comparisons, phylogenetic topologies and intron characterizations, each highly expanded ABC transporter family of T. thermophila was classified into several distinct groups, and hypotheses about their evolutionary relationships are presented. A comprehensive microarray analysis revealed divergent expression patterns among the members of the ABC transporter superfamily during different states of physiology and development. Many of the relatively recently formed duplicate pairs within individual ABC transporter families exhibit significantly different expression patterns. Further analysis showed that multiple mechanisms have led to functional divergence that is responsible for the preservation of duplicated genes. Conclusion Gene duplications have resulted in an extensive expansion of the superfamily of ABC transporters in the Tetrahymena genome, making it the largest example of its kind reported in any organism to date. Multiple independent duplications and subsequent divergence contributed to the formation of different families of ABC transporter genes. Many of the members within a gene family exhibit different expression patterns. The combination of gene duplication followed by both sequence divergence and acquisition of new patterns of

  18. Transportomics: screening for substrates of ABC transporters in body fluids using vesicular transport assays

    NARCIS (Netherlands)

    Krumpochova, Petra; Sapthu, Sunny; Brouwers, Jos F.; de Haas, Marcel; de Vos, Ric; Borst, Piet; van de Wetering, Koen

    2012-01-01

    The ATP-binding cassette (ABC) genes encode the largest family of transmembrane proteins. ABC transporters translocate a wide variety of substrates across membranes, but their physiological function is often incompletely understood. We describe a new method to study the substrate spectrum of ABC

  19. A novel ATP-binding cassette transporter is responsible for resistance to viologen herbicides in the cyanobacterium Synechocystis sp. PCC 6803

    Czech Academy of Sciences Publication Activity Database

    Prosecká, J.; Orlov, V. N.; Fantin, Y. S.; Zinchenko, V. V.; Babykin, M. M.; Tichý, Martin

    2009-01-01

    Roč. 276, č. 15 (2009), s. 4001-4011 ISSN 1742-464X R&D Projects: GA MŠk ME 881 Institutional research plan: CEZ:AV0Z50200510 Keywords : ABC-type transporter * cyanobacteria * oxidative stress Subject RIV: EE - Microbiology, Virology Impact factor: 3.042, year: 2009

  20. Hematopoietic stem cells exhibit a specific ABC transporter gene expression profile clearly distinct from other stem cells.

    NARCIS (Netherlands)

    Tang, L.; Bergevoet, S.M.; Gilissen, C.F.H.A.; Witte, T.J.M. de; Jansen, J.H.; Reijden, B.A. van der; Raymakers, R.A.P.

    2010-01-01

    BACKGROUND: ATP-binding cassette (ABC) transporters protect cells against unrelated (toxic) substances by pumping them across cell membranes. Earlier we showed that many ABC transporters are highly expressed in hematopoietic stem cells (HSCs) compared to more committed progenitor cells. The ABC

  1. ABC and MFS transporters from Botrytis cinerea involved in sensitivity to fungicides and natural toxic compounds

    NARCIS (Netherlands)

    Hayashi, K.

    2003-01-01

    ATP-binding cassette (ABC) and major facilitator superfamily (MFS) transporters are two major classes of proteins involved in drug resistance. ABC transporter proteins are primary transporters that use the energy generated by ATP hydrolysis to transport drugs over membranes, while MFS transport

  2. Transportomics: screening for substrates of ABC transporters in body fluids using vesicular transport assays.

    NARCIS (Netherlands)

    Krumpochova, P; Sapthu, S.; Brouwers, J.F.H.M.; de Haas, M.; de Vos, R.; Borst, P.; van de Wetering, K.

    2013-01-01

    ABSTRACT The ATP-binding cassette (ABC) genes encode the largest family of transmembrane proteins. ABC transporters translocate a wide variety of substrates across membranes, but their physiological function is often incompletely understood. We describe a new method to study the substrate spectrum

  3. Energy Coupling Factor-Type ABC Transporters for Vitamin Uptake in Prokaryotes

    NARCIS (Netherlands)

    Erkens, Guus B.; Dosz-Majsnerowska, Maria; ter Beek, Josy; Slotboom, Dirk Jan

    2012-01-01

    Energy coupling factor (ECF) transporters are a subgroup of ATP-binding cassette (ABC) transporters involved in the uptake of vitamins and micronutrients in prokaryotes. In contrast to classical ABC importers, ECF transporters do not make use of water-soluble substrate binding proteins or domains

  4. Structural and mechanistic insights into ABC-type ECF transporters for vitamin uptake

    NARCIS (Netherlands)

    Dosz-Majsnerowska, Maria

    2014-01-01

    Dit proefschrift gaat over de relatie tussen de structuur en het mechanisme van ABC-type ECF transporters voor vitamines, uit de bacterie Lactococcus lactis. Energy-Coupling Factor (ECF) transporters vormen een subgroep van de ATP-binding cassette (ABC) transporters en zijn betrokken bij de opname

  5. Structural diversity of ABC transporters

    Science.gov (United States)

    ter Beek, Josy; Guskov, Albert

    2014-01-01

    ATP-binding cassette (ABC) transporters form a large superfamily of ATP-dependent protein complexes that mediate transport of a vast array of substrates across membranes. The 14 currently available structures of ABC transporters have greatly advanced insight into the transport mechanism and revealed a tremendous structural diversity. Whereas the domains that hydrolyze ATP are structurally related in all ABC transporters, the membrane-embedded domains, where the substrates are translocated, adopt four different unrelated folds. Here, we review the structural characteristics of ABC transporters and discuss the implications of this structural diversity for mechanistic diversity. PMID:24638992

  6. Modulation of microRNA Expression in Subjects with Metabolic Syndrome and Decrease of Cholesterol Efflux from Macrophages via microRNA-33-Mediated Attenuation of ATP-Binding Cassette Transporter A1 Expression by Statins.

    Science.gov (United States)

    Chen, Wei-Ming; Sheu, Wayne H-H; Tseng, Pei-Chi; Lee, Tzong-Shyuan; Lee, Wen-Jane; Chang, Pey-Jium; Chiang, An-Na

    2016-01-01

    Metabolic syndrome (MetS) is a complicated health problem that encompasses a variety of metabolic disorders. In this study, we analyzed the relationship between the major biochemical parameters associated with MetS and circulating levels of microRNA (miR)-33, miR-103, and miR-155. We found that miRNA-33 levels were positively correlated with levels of fasting blood glucose, glycosylated hemoglobin A1c, total cholesterol, LDL-cholesterol, and triacylglycerol, but negatively correlated with HDL-cholesterol levels. In the cellular study, miR-33 levels were increased in macrophages treated with high glucose and cholesterol-lowering drugs atorvastatin and pitavastatin. miR-33 has been reported to play an essential role in cholesterol homeostasis through ATP-binding cassette transporter A1 (ABCA1) regulation and reverse cholesterol transport. However, the molecular mechanism underlying the linkage between miR-33 and statin treatment remains unclear. In the present study, we investigated whether atorvastatin and pitavastatin exert their functions through the modulation of miR-33 and ABCA1-mediated cholesterol efflux from macrophages. The results showed that treatment of the statins up-regulated miR-33 expression, but down-regulated ABCA1 mRNA levels in RAW264.7 cells and bone marrow-derived macrophages. Statin-mediated ABCA1 regulation occurs at the post-transcriptional level through targeting of the 3'-UTR of the ABCA1 transcript by miR-33. Additionally, we found significant down-regulation of ABCA1 protein expression in macrophages treated with statins. Finally, we showed that high glucose and statin treatment significantly suppressed cholesterol efflux from macrophages. These findings have highlighted the complexity of statins, which may exert detrimental effects on metabolic abnormalities through regulation of miR-33 target genes.

  7. Modulation of microRNA Expression in Subjects with Metabolic Syndrome and Decrease of Cholesterol Efflux from Macrophages via microRNA-33-Mediated Attenuation of ATP-Binding Cassette Transporter A1 Expression by Statins.

    Directory of Open Access Journals (Sweden)

    Wei-Ming Chen

    Full Text Available Metabolic syndrome (MetS is a complicated health problem that encompasses a variety of metabolic disorders. In this study, we analyzed the relationship between the major biochemical parameters associated with MetS and circulating levels of microRNA (miR-33, miR-103, and miR-155. We found that miRNA-33 levels were positively correlated with levels of fasting blood glucose, glycosylated hemoglobin A1c, total cholesterol, LDL-cholesterol, and triacylglycerol, but negatively correlated with HDL-cholesterol levels. In the cellular study, miR-33 levels were increased in macrophages treated with high glucose and cholesterol-lowering drugs atorvastatin and pitavastatin. miR-33 has been reported to play an essential role in cholesterol homeostasis through ATP-binding cassette transporter A1 (ABCA1 regulation and reverse cholesterol transport. However, the molecular mechanism underlying the linkage between miR-33 and statin treatment remains unclear. In the present study, we investigated whether atorvastatin and pitavastatin exert their functions through the modulation of miR-33 and ABCA1-mediated cholesterol efflux from macrophages. The results showed that treatment of the statins up-regulated miR-33 expression, but down-regulated ABCA1 mRNA levels in RAW264.7 cells and bone marrow-derived macrophages. Statin-mediated ABCA1 regulation occurs at the post-transcriptional level through targeting of the 3'-UTR of the ABCA1 transcript by miR-33. Additionally, we found significant down-regulation of ABCA1 protein expression in macrophages treated with statins. Finally, we showed that high glucose and statin treatment significantly suppressed cholesterol efflux from macrophages. These findings have highlighted the complexity of statins, which may exert detrimental effects on metabolic abnormalities through regulation of miR-33 target genes.

  8. Brucella abortus mutants lacking ATP-binding cassette transporter proteins are highly attenuated in virulence and confer protective immunity against virulent B. abortus challenge in BALB/c mice.

    Science.gov (United States)

    Truong, Quang Lam; Cho, Youngjae; Park, Soyeon; Park, Bo-Kyoung; Hahn, Tae-Wook

    2016-06-01

    Brucella abortus RB51 is an attenuated vaccine strain that has been most frequently used for bovine brucellosis. Although it is known to provide good protection in cattle, it still has some drawbacks including resistance to rifampicin, residual virulence and pathogenicity in humans. Thus, there has been a continuous interest on new safe and effective bovine vaccine candidates. In the present study, we have constructed unmarked mutants by deleting singly cydD and cydC genes, which encode ATP-binding cassette transporter proteins, from the chromosome of the virulent Brucella abortus isolate from Korean cow (referred to as IVK15). Both IVK15ΔcydD and ΔcydC mutants showed increased sensitivity to metal ions, hydrogen peroxide and acidic pH, which are mimic to intracellular environment during host infection. Additionally, the mutants exhibited a significant growth defect in RAW264.7 cells and greatly attenuated in mice. Vaccination of mice with either IVK15ΔcydC or IVK15ΔcydD mutant could elicit an anti-Brucella specific immunoglobulin G (IgG) and IgG subclass responses as well as enhance the secretion of interferon-gamma, and provided better protection against challenge with B. abortus strain 2308 than with the commercial B. abortus strain RB51 vaccine. Collectively, these results suggest that both IVK15ΔcydC and IVK15ΔcydD mutants could be an attenuated vaccine candidate against B. abortus. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. ABC transporter genes and risk of type 2 diabetes

    DEFF Research Database (Denmark)

    Schou, Jesper; Tybjærg-Hansen, Anne; Møller, Holger Jon

    2012-01-01

    Alterations of pancreatic β-cell cholesterol content may contribute to β-cell dysfunction. Two important determinants of intracellular cholesterol content are the ATP-binding cassette (ABC) transporters A1 (ABCA1) and -G1 (ABCG1). Whether genetic variation in ABCA1 and ABCG1 predicts risk of type 2...... diabetes in the general population is unknown....

  10. ABC transporters from Botrytis cinerea in biotic and abiotic interactions

    NARCIS (Netherlands)

    Schoonbeek, H.

    2004-01-01

    Botrytis cinereais the causal agent of grey mould disease on a wide variety of crop plants. It is relatively insensitive to natural and synthetic fungitoxic compounds. This thesis describes how ABC (ATP-binding cassette) transporters contribute to protection by actively

  11. Functional analysis of candidate ABC transporter proteins for sitosterol transport

    DEFF Research Database (Denmark)

    Albrecht, C; Elliott, J I; Sardini, A

    2002-01-01

    Two ATP-binding cassette (ABC) proteins, ABCG5 and ABCG8, have recently been associated with the accumulation of dietary cholesterol in the sterol storage disease sitosterolemia. These two 'half-transporters' are assumed to dimerize to form the complete sitosterol transporter which reduces the ab...

  12. Genetic variant of V825I in the ATP-binding cassette transporter A1 gene and serum lipid levels in the Guangxi Bai Ku Yao and Han populations.

    Science.gov (United States)

    Cao, Xiao-Li; Yin, Rui-Xing; Wu, Dong-Feng; Miao, Lin; Aung, Lynn Htet Htet; Hu, Xi-Jiang; Li, Qing; Yan, Ting-Ting; Lin, Wei-Xiong; Pan, Shang-Ling

    2011-01-19

    Several genetic variants in the ATP-binding cassette transporter A1 (ABCA1) gene have associated with modifications of serum high-density lipoprotein cholesterol (HDL-C) levels and the susceptibility for coronary heart disease, but the findings are still controversial in diverse racial/ethnic groups. Bai Ku Yao is an isolated subgroup of the Yao minority in southern China. The present study was undertaken to detect the possible association of V825I (rs2066715) polymorphism in the ABCA1 gene and several environmental factors with serum lipid levels in the Guangxi Bai Ku Yao and Han populations. A total of 677 subjects of Bai Ku Yao and 646 participants of Han Chinese were randomly selected from our previous stratified randomized cluster samples. Polymerase chain reaction and restriction fragment length polymorphism assay combined with gel electrophoresis were performed for the genotyping of V825I variant, and then confirmed by direct sequencing. The levels of serum total cholesterol (TC), HDL-C, apolipoprotein (Apo) AI and ApoB were lower in Bai Ku Yao than in Han (P 0.05); respectively. The frequency of GG, GA and AA genotypes was 33.7%, 47.4% and 18.9% in Bai Ku Yao, and 33.4%, 48.6% and 18.0% in Han (P > 0.05); respectively. There was no difference in the genotypic and allelic frequencies between males and females in the both ethnic groups. The subjects with AA genotype in Bai Ku Yao had higher serum TC levels than the subjects with GG and GA genotypes (P blood pressure in both ethnic groups (P < 0.05-0.001). The present study suggests that the V825I polymorphism in the ABCA1 gene is associated with male serum HDL-C and ApoAI levels in the Han, and serum TC levels in the Bai Ku Yao populations. The difference in the association of V825I polymorphism and serum lipid levels between the two ethnic groups might partly result from different ABCA1 gene-environmental interactions.

  13. Genetic variant of V825I in the ATP-binding cassette transporter A1 gene and serum lipid levels in the Guangxi Bai Ku Yao and Han populations

    Directory of Open Access Journals (Sweden)

    Yan Ting-Ting

    2011-01-01

    Full Text Available Abstract Background Several genetic variants in the ATP-binding cassette transporter A1 (ABCA1 gene have associated with modifications of serum high-density lipoprotein cholesterol (HDL-C levels and the susceptibility for coronary heart disease, but the findings are still controversial in diverse racial/ethnic groups. Bai Ku Yao is an isolated subgroup of the Yao minority in southern China. The present study was undertaken to detect the possible association of V825I (rs2066715 polymorphism in the ABCA1 gene and several environmental factors with serum lipid levels in the Guangxi Bai Ku Yao and Han populations. Methods A total of 677 subjects of Bai Ku Yao and 646 participants of Han Chinese were randomly selected from our previous stratified randomized cluster samples. Polymerase chain reaction and restriction fragment length polymorphism assay combined with gel electrophoresis were performed for the genotyping of V825I variant, and then confirmed by direct sequencing. Results The levels of serum total cholesterol (TC, HDL-C, apolipoprotein (Apo AI and ApoB were lower in Bai Ku Yao than in Han (P P > 0.05; respectively. The frequency of GG, GA and AA genotypes was 33.7%, 47.4% and 18.9% in Bai Ku Yao, and 33.4%, 48.6% and 18.0% in Han (P > 0.05; respectively. There was no difference in the genotypic and allelic frequencies between males and females in the both ethnic groups. The subjects with AA genotype in Bai Ku Yao had higher serum TC levels than the subjects with GG and GA genotypes (P P P P Conclusion The present study suggests that the V825I polymorphism in the ABCA1 gene is associated with male serum HDL-C and ApoAI levels in the Han, and serum TC levels in the Bai Ku Yao populations. The difference in the association of V825I polymorphism and serum lipid levels between the two ethnic groups might partly result from different ABCA1 gene-enviromental interactions.

  14. Plant ABC transporters enable many unique aspects of a terrestrial plant's lifestyle

    DEFF Research Database (Denmark)

    Hwang, Jae-Ung; Song, Won-Yong; Hong, Daewoong

    2016-01-01

    Terrestrial plants have two to four times more ATP-binding cassette (ABC) transporter genes than other organisms, including their ancestral microalgae. Recent studies found that plants harboring mutations in these transporters exhibit dramatic phenotypes, many of which are related to developmental...

  15. Transcriptome-based identification of ABC transporters in the western tarnished plant bug lygus hesperus

    Science.gov (United States)

    ATP-binding cassette (ABC) transporters are a large superfamily of proteins that mediate diverse physiological functions by coupling ATP hydrolysis with substrate transport across lipid membranes. In insects, these proteins play roles in metabolism, development, eye pigmentation, and xenobiotic cle...

  16. Functional Diversity of Tandem Substrate-Binding Domains in ABC Transporters from Pathogenic Bacteria

    NARCIS (Netherlands)

    Fulyani, Faizah; Schuurman-Wolters, Gea K.; Vujicic - Zagar, Andreja; Guskov, Albert; Slotboom, Dirk-Jan; Poolman, Bert

    2013-01-01

    The ATP-binding cassette (ABC) transporter GInPQ is an essential uptake system for amino acids in gram-positive pathogens and related nonpathogenic bacteria. The transporter has tandem substrate-binding domains (SBDs) fused to each transmembrane domain, giving rise to four SBDs per functional

  17. Human and Rat ABC Transporter Efflux of Bisphenol A and Bisphenol A Glucuronide: Interspecies Comparison and Implications for Pharmacokinetic Assessment

    Science.gov (United States)

    Significant interspecies differences exist between human and rodent with respect to absorption, distribution, and excretion of bisphenol A (BPA) and its primary metabolite, BPA-glucuronide (BPA-G). ATP-Binding Cassette (ABC) transporter enzymes play important roles in these physi...

  18. Functional analysis of ABC transporter genes from Botrytis cinerea identifies BcatrB as a transporter of eugenol

    NARCIS (Netherlands)

    Schoonbeek, H.; Nistelrooy, van J.G.M.; Waard, de M.A.

    2003-01-01

    The role of multiple ATP-binding cassette (ABC) and major facilitator superfamily (MFS) transporter genes from the plant pathogenic fungus Botrytis cinerea in protection against natural fungitoxic compounds was studied by expression analysis and phenotyping of gene-replacement mutants. The

  19. Equilibrated atomic models of outward-facing P-glycoprotein and effect of ATP binding on structural dynamics.

    Science.gov (United States)

    Pan, Lurong; Aller, Stephen G

    2015-01-20

    P-glycoprotein (Pgp) is an ATP-binding cassette (ABC) transporter that alternates between inward- and outward-facing conformations to capture and force substrates out of cells like a peristaltic pump. The high degree of similarity in outward-facing structures across evolution of ABC transporters allowed construction of a high-confidence outward-facing Pgp atomic model based on crystal structures of outward-facing Sav1866 and inward-facing Pgp. The model adhered to previous experimentally determined secondary- and tertiary- configurations during all-atom molecular dynamics simulations in the presence or absence of MgATP. Three long lasting (>100 ns) meta-stable states were apparent in the presence of MgATP revealing new insights into alternating access. The two ATP-binding pockets are highly asymmetric resulting in differential control of overall structural dynamics and allosteric regulation of the drug-binding pocket. Equilibrated Pgp has a considerably different electrostatic profile compared to Sav1866 that implicates significant kinetic and thermodynamic differences in transport mechanisms.

  20. Lantibiotic transporter requires cooperative functioning of the peptidase domain and the ATP binding domain.

    Science.gov (United States)

    Nishie, Mami; Sasaki, Makoto; Nagao, Jun-ichi; Zendo, Takeshi; Nakayama, Jiro; Sonomoto, Kenji

    2011-04-01

    Lantibiotics are ribosomally synthesized and post-translationally modified peptide antibiotics that contain unusual amino acids such as dehydro and lanthionine residues. Nukacin ISK-1 is a class II lantibiotic, whose precursor peptide (NukA) is modified by NukM to form modified NukA. ATP-binding cassette (ABC) transporter NukT is predicted to cleave off the N-terminal leader peptide of modified NukA and secrete the mature peptide. Multiple sequence alignments revealed that NukT has an N-terminal peptidase domain (PEP) and a C-terminal ATP binding domain (ABD). Previously, in vitro reconstitution of NukT has revealed that NukT peptidase activity depends on ATP hydrolysis. Here, we constructed a series of NukT mutants and investigated their transport activity in vivo and peptidase activity in vitro. Most of the mutations of the conserved residues of PEP or ABD resulted in failure of nukacin ISK-1 production and accumulation of modified NukA inside the cells. NukT(N106D) was found to be the only mutant capable of producing nukacin ISK-1. Asn(106) is conserved as Asp in other related ABC transporters. Additionally, an in vitro peptidase assay of NukT mutants demonstrated that PEP is on the cytosolic side and all of the ABD mutants as well as PEP (with the exception of NukT(N106D)) did not have peptidase activity in vitro. Taken together, these observations suggest that the leader peptide is cleaved off inside the cells before peptide secretion; both PEP and ABD are important for NukT peptidase activity, and cooperation between these two domains inside the cells is indispensable for proper functioning of NukT.

  1. Lantibiotic Transporter Requires Cooperative Functioning of the Peptidase Domain and the ATP Binding Domain*

    Science.gov (United States)

    Nishie, Mami; Sasaki, Makoto; Nagao, Jun-ichi; Zendo, Takeshi; Nakayama, Jiro; Sonomoto, Kenji

    2011-01-01

    Lantibiotics are ribosomally synthesized and post-translationally modified peptide antibiotics that contain unusual amino acids such as dehydro and lanthionine residues. Nukacin ISK-1 is a class II lantibiotic, whose precursor peptide (NukA) is modified by NukM to form modified NukA. ATP-binding cassette (ABC) transporter NukT is predicted to cleave off the N-terminal leader peptide of modified NukA and secrete the mature peptide. Multiple sequence alignments revealed that NukT has an N-terminal peptidase domain (PEP) and a C-terminal ATP binding domain (ABD). Previously, in vitro reconstitution of NukT has revealed that NukT peptidase activity depends on ATP hydrolysis. Here, we constructed a series of NukT mutants and investigated their transport activity in vivo and peptidase activity in vitro. Most of the mutations of the conserved residues of PEP or ABD resulted in failure of nukacin ISK-1 production and accumulation of modified NukA inside the cells. NukT(N106D) was found to be the only mutant capable of producing nukacin ISK-1. Asn106 is conserved as Asp in other related ABC transporters. Additionally, an in vitro peptidase assay of NukT mutants demonstrated that PEP is on the cytosolic side and all of the ABD mutants as well as PEP (with the exception of NukT(N106D)) did not have peptidase activity in vitro. Taken together, these observations suggest that the leader peptide is cleaved off inside the cells before peptide secretion; both PEP and ABD are important for NukT peptidase activity, and cooperation between these two domains inside the cells is indispensable for proper functioning of NukT. PMID:21303905

  2. Novel understanding of ABC transporters ABCB1/MDR/P-glycoprotein, ABCC2/MRP2, and ABCG2/BCRP in colorectal pathophysiology

    DEFF Research Database (Denmark)

    Andersen, Vibeke; Svenningsen, Katrine; Almind Knudsen, Lina

    2015-01-01

    AIM: To evaluate ATP-binding cassette (ABC) transporters in colonic pathophysiology as they had recently been related to colorectal cancer (CRC) development. METHODS: Literature search was conducted on PubMed using combinations of the following terms: ABC transporters, ATP binding cassette....../Mdr1a, abcc2/Mrp2, abcg2/Bcrp, knock-out mice, tight junction, membrane lipid function. RESULTS: Recently, human studies reported that changes in the levels of ABC transporters were early events in the adenoma-carcinoma sequence leading to CRC. A link between ABCB1, high fat diet and gut microbes...... translocation from one side to the other of the cell membrane lipid bilayer by ABC transporters affecting inflammatory response and/or function of tight junctions, phagocytosis and vesicle trafficking. Also, diet and microbes give rise to molecules which are potential substrates for the ABC transporters...

  3. The role of ABC proteins Aus1p and Pdr11p in the uptake of external sterols in yeast: dehydroergosterol fluorescence study

    DEFF Research Database (Denmark)

    Kohut, Peter; Wüstner, Daniel; Hronska, L

    2011-01-01

    of sterol molecules into plasma membrane is not spontaneous but requires assistance of two ABC (ATP-binding cassette) pumps--Aus1p or Pdr11p. DHE taken up by uptake-competent hem1ΔAUS1PDR11 cells could be directly visualized by UV-sensitive wide field fluorescence microscopy. HPLC analysis of sterols...

  4. Conservation of targeting but divergence in function and quality control of peroxisomal ABC transporters: an analysis using cross-kingdom expression

    NARCIS (Netherlands)

    Zhang, Xuebin; de Marcos Lousa, Carine; Schutte-Lensink, Nellie; Ofman, Rob; Wanders, Ronald J.; Baldwin, Stephen A.; Baker, Alison; Kemp, Stephan; Theodoulou, Frederica L.

    2011-01-01

    ABC (ATP-binding cassette) subfamily D transporters are found in all eukaryotic kingdoms and are known to play essential roles in mammals and plants; however, their number, organization and physiological contexts differ. Via cross-kingdom expression experiments, we have explored the conservation of

  5. ABC transporters in the CNS - an inventory.

    Science.gov (United States)

    Hartz, A M S; Bauer, B

    2011-04-01

    In the present review we provide a summary of ATP-binding cassette (ABC) transporters in the central nervous system (CNS). Our review is focused on transporters of the ABC A, B, C, D, and G families that have been detected in the cells of the neurovascular unit/blood-brain barrier including brain capillary endothelial cells, pericytes, astrocytes, and neurons, as well as in other brain cells, such as microglia, oligodendrocytes, and choroid plexus epithelial cells. In this review, we provide an overview, organized by ABC family, of transporter expression, localization, and function. We summarize recent findings on ABC transporter regulation in the CNS and address the role of ABC transporters in CNS diseases including brain cancer, seizures/epilepsy, and Alzheimer's disease. Finally, we discuss new therapeutic strategies focused on ABC transporters in CNS disease.

  6. An inventory of the human ABC proteins.

    Science.gov (United States)

    Klein, I; Sarkadi, B; Váradi, A

    1999-12-06

    Currently 30 human ABC proteins are represented by full sequences in various databases, and this paper provides a brief overview of these proteins. ABC proteins are composed of transmembrane domains (TMDs), and nucleotide binding domains (NBDs, or ATP-binding cassettes, ABSs). The arrangement of these domains, together with available membrane topology models of the family members, are presented. Based on their sequence similarity scores, the members of the human ABC protein family can be grouped into eight subfamilies. At present the MDR/TAP, the ALD, the MRP/CFTR, the ABC1, the White, the RNAseL inhibitor, the ANSA, and the GCN20 subfamilies are identified. Mutations of many human ABC proteins are known to be causative in inherited diseases, and a short description of the molecular pathology of these ABC gene-related genetic diseases is also provided.

  7. Interaction of BDE-47 and its Hydroxylated Metabolite 6-OH-BDE-47 with the Human ABC Efflux Transporters P-gp and BCRP: Considerations for Human Exposure and Risk Assessment

    Science.gov (United States)

    ATP binding cassette (ABC) transporters, including P-glycoprotein (P-gp; also known as MDR1, ABCB1) and breast cancer resistance protein (BCRP; also known as ABCG2), are membrane-bound proteins that mediate the cellular efflux of xenobiotics as an important defense against chemic...

  8. An ATP Binding Cassette Transporter Mediates the Uptake of α-(1,6)-Linked Dietary Oligosaccharides in Bifidobacterium and Correlates with Competitive Growth on These Substrates

    DEFF Research Database (Denmark)

    Hansen, Morten Ejby; Fredslund, Folmer; Andersen, Joakim Mark

    2016-01-01

    composition with preference for the trisaccharides raffinose and panose. This preference is also reflected in the -(1,6)-galactoside uptake profile of the bacterium. Structures of BlG16BP in complex with raffinose and panose revealed the basis for the remarkable ligand binding plasticity of BlG16BP, which...... of raffinose provides an important competitive advantage, particularly against dominant Bacteroides that lack glycan-specific ABC-transporters. This novel insight highlights the role of glycan transport in defining the metabolic specialization of gut bacteria....

  9. Split tasks of asymmetric nucleotide-binding sites in the heterodimeric ABC exporter EfrCD.

    Science.gov (United States)

    Hürlimann, Lea M; Hohl, Michael; Seeger, Markus A

    2017-06-01

    Many heterodimeric ATP-binding cassette (ABC) exporters evolved asymmetric ATP-binding sites containing a degenerate site incapable of ATP hydrolysis due to noncanonical substitutions in conserved sequence motifs. Recent studies revealed that nucleotide binding to the degenerate site stabilizes contacts between the nucleotide-binding domains (NBDs) of the inward-facing transporter and regulates ATP hydrolysis at the consensus site via allosteric coupling mediated by the D-loops. However, it is unclear whether nucleotide binding to the degenerate site is strictly required for substrate transport. In this study, we examined the functional consequences of a systematic set of mutations introduced at the degenerate and consensus site of the multidrug efflux pump EfrCD of Enterococcus faecalis. Mutating motifs which differ among the two ATP-binding sites (Walker B, switch loop, and ABC signature) or which are involved in interdomain communication (D-loop and Q-loop) led to asymmetric results in the functional assays and were better tolerated at the degenerate site. This highlights the importance of the degenerate site to allosterically regulate the events at the consensus site. Mutating invariant motifs involved in ATP binding and NBD closure (A-loop and Walker A) resulted in equally reduced transport activities, regardless at which ATP-binding site they were introduced. In contrast to previously investigated heterodimeric ABC exporters, mutation of the degenerate site Walker A lysine completely inactivated ATPase activity and substrate transport, indicating that ATP binding to the degenerate site is essential for EfrCD. This study provides novel insights into the split tasks of asymmetric ATP-binding sites of heterodimeric ABC exporters. © 2017 The Authors. The FEBS Journal published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.

  10. A putative ABC transporter is involved in negative regulation of biofilm formation by Listeria monocytogenes

    DEFF Research Database (Denmark)

    Zhu, Xinna; Long, Fei; Chen, Yonghui

    2008-01-01

    with enhanced ability of biofilm-formation generated via transposon Tn917 mutagenesis of L. monocytogenes 4b G. In this mutant, a Tn917 insertion has disrupted the coding region of the gene encoding a putative ATP binding cassette (ABC) transporter permease identical to Lmof2365_1771 (a putative ABC...... regulation of biofilm formation by L. monocytogenes 4b G. The immediate gene upstream of lm.G_1771 encoded an ATP-binding protein. Bioinformatics analysis suggested that these two genes were organized into an operon and that their proteins formed an export ABC transporter. Here, we report...... the characterization of the mutant and identification of a novel ABC transporter that functions in negative regulation of biofilm formation in L. monocytogenes....

  11. Yeast ABC proteins involved in multidrug resistance.

    Science.gov (United States)

    Piecuch, Agata; Obłąk, Ewa

    2014-03-01

    Pleiotropic drug resistance is a complex phenomenon that involves many proteins that together create a network. One of the common mechanisms of multidrug resistance in eukaryotic cells is the active efflux of a broad range of xenobiotics through ATP-binding cassette (ABC) transporters. Saccharomyces cerevisiae is often used as a model to study such activity because of the functional and structural similarities of its ABC transporters to mammalian ones. Numerous ABC transporters are found in humans and some are associated with the resistance of tumors to chemotherapeutics. Efflux pump modulators that change the activity of ABC proteins are the most promising candidate drugs to overcome such resistance. These modulators can be chemically synthesized or isolated from natural sources (e.g., plant alkaloids) and might also be used in the treatment of fungal infections. There are several generations of synthetic modulators that differ in specificity, toxicity and effectiveness, and are often used for other clinical effects.

  12. Expression and regulation of prostaglandin transporters, ATP-binding cassette, subfamily C, member 1 and 9, and solute carrier organic anion transporter family, member 2A1 and 5A1 in the uterine endometrium during the estrous cycle and pregnancy in pigs

    Directory of Open Access Journals (Sweden)

    Hwanhee Jang

    2017-05-01

    Full Text Available Objective Prostaglandins (PGs function in various reproductive processes, including luteolysis, maternal pregnancy recognition, conceptus development, and parturition. Our earlier study has shown that PG transporters ATP-binding cassette, subfamily C, member 4 (ABCC4 and solute carrier organic anion transporter family, member 2A1 (SLCO2A1 are expressed in the uterine endometrium in pigs. Since several other PG transporters such as ABCC1, ABCC9, SLCO4C1, and SLCO5A1 are known to be present in the uterine endometrium, this study investigated the expression of these PG transporters in the porcine uterine endometrium and placenta. Methods Uterine endometrial tissues were obtained from gilts on day (D 12 and D15 of the estrous cycle and days 12, 15, 30, 60, 90, and 114 of pregnancy. Results ABCC1, ABCC9, SLCO4C1, and SLCO5A1 mRNAs were expressed in the uterine endometrium, and levels of expression changed during the estrous cycle and pregnancy. Expression of ABCC1 and ABCC9 mRNAs was localized mainly to luminal and glandular epithelial cells in the uterine endometrium, and chorionic epithelial cells during pregnancy. Conceptuses during early pregnancy and chorioallantoic tissues from mid to late pregnancy also expressed these PG transporters. Estradiol-17β increased the expression of ABCC1 and SLCO5A1, but not ABCC9 and SLCO4C1 mRNAs and increasing doses of interleukin-1β induced the expression of ABCC9, SLCO4C1, and SLCO5A1 mRNAs in endometrial explant tissues. Conclusion These data showed that several PG transporters such as ABCC1, ABCC9, SLCO4C1, and SLCO5A1 were expressed at the maternal-conceptus interface, suggesting that these PG transporters may play an important role in the establishment and maintenance of pregnancy by regulating PG transport in the uterine endometrium and placenta in pigs.

  13. ATP-Binding Cassette Transporters, Atherosclerosis, and Inflammation

    NARCIS (Netherlands)

    Westerterp, Marit; Bochem, Andrea E.; Yvan-Charvet, Laurent; Murphy, Andrew J.; Wang, Nan; Tall, Alan R.

    2014-01-01

    Although recent genome-wide association studies have called into question the causal relationship between high-density lipoprotein (HDL) cholesterol levels and cardiovascular disease, ongoing research in animals and cells has produced increasing evidence that cholesterol efflux pathways mediated by

  14. Bioinformatic survey of ABC transporters in dermatophytes.

    Science.gov (United States)

    Gadzalski, Marek; Ciesielska, Anita; Stączek, Paweł

    2016-01-15

    ATP binding cassette (ABC) transporters constitute a very large and ubiquitous superfamily of membrane proteins. They are responsible for ATP hydrolysis driven translocation of countless substrates. Being a very old and diverse group of proteins present in all organisms they share a common feature, which is the presence of an evolutionary conservative nucleotide binding domain (NBD)--the engine that drives the transport. Another common domain is a transmembrane domain (TMD) which consists of several membrane-spanning helices. This part of protein is substrate-specific, thus it is much more variable. ABC transporters are known for driving drug efflux in many pathogens and cancer cells, therefore they are the subject of extensive studies. There are many examples of conferring a drug resistance phenotype in fungal pathogens by ABC transporters, however, little is known about these proteins in dermatophytes--a group of fungi causing superficial mycoses. So far only a single ABC transporter has been extensively studied in this group of pathogens. We analyzed available genomic sequences of seven dermatophyte species in order to provide an insight into dermatophyte ABC protein inventory. Phylogenetic studies of ABC transporter genes and their products were conducted and included ABC transporters of other fungi. Our results show that each dermatophyte genome studied possesses a great variety of ABC transporter genes. Detailed analysis of selected genes and their products indicates that relatively recent duplication of ABC transporter genes could lead to novel substrate specificity. Copyright © 2015 Elsevier B.V. All rights reserved.

  15. ABC proteins protect the human body and maintain optimal health.

    Science.gov (United States)

    Ueda, Kazumitsu

    2011-01-01

    Human MDR1, a multi-drug transporter gene, was isolated as the first of the eukaryote ATP Binding Cassette (ABC) proteins from a multidrug-resistant carcinoma cell line in 1986. To date, over 25 years, many ABC proteins have been found to play important physiological roles by transporting hydrophobic compounds. Defects in their functions cause various diseases, indicating that endogenous hydrophobic compounds, as well as water-soluble compounds, are properly transported by transmembrane proteins. MDR1 transports a large number of structurally unrelated drugs and is involved in their pharmacokinetics, and thus is a key factor in drug interaction. ABCA1, an ABC protein, eliminates excess cholesterol in peripheral cells by generating HDL. Because ABCA1 is a key molecule in cholesterol homeostasis, its function and expression are highly regulated. Eukaryote ABC proteins function on the body surface facing the outside and in organ pathways to adapt to the extracellular environment and protect the body to maintain optimal health.

  16. Functional expression and characterization of plant ABC transporters in Xenopus laevis oocytes for transport engineering purposes

    DEFF Research Database (Denmark)

    Xu, Deyang; Veres, Dorottya; Belew, Zeinu Mussa

    2016-01-01

    Transport engineering in bioengineering is aimed at efficient export of the final product to reduce toxicity and feedback inhibition and to increase yield. The ATP-binding cassette (ABC) transporters with their highly diverse substrate specificity and role in cellular efflux are potentially...... suitable in transport engineering approaches, although their size and high number of introns make them notoriously difficult to clone. Here, we report a novel in planta “exon engineering” strategy for cloning of full-length coding sequence of ABC transporters followed by methods for biochemical...... provided will hopefully contribute to more successful transport engineering in synthetic biology....

  17. The multidrug ABC transporter BmrC/BmrD of Bacillus subtilis is regulated via a ribosome-mediated transcriptional attenuation mechanism

    OpenAIRE

    Reilman, E.; Mars, R. A. T.; van Dijl, J. M.; Denham, Emma

    2014-01-01

    Expression of particular drug transporters in response to antibiotic pressure is a critical element in the development of bacterial multidrug resistance, and represents a serious concern for human health. To obtain a better understanding of underlying regulatory mechanisms, we have dissected the transcriptional activation of the ATP-binding cassette (ABC) transporter BmrC/BmrD of the Gram-positive model bacterium Bacillus subtilis. By using promoter-GFP fusions and live cell array technology,...

  18. Inventory and function of yeast ABC proteins: about sex, stress, pleiotropic drug and heavy metal resistance.

    Science.gov (United States)

    Bauer, B E; Wolfger, H; Kuchler, K

    1999-12-06

    Saccharomyces cerevisiae was the first eukaryotic organism whose complete genome sequence has been determined, uncovering the existence of numerous genes encoding proteins of the ATP-binding cassette (ABC) family. Fungal ABC proteins are implicated in a variety of cellular functions, ranging from clinical drug resistance development, pheromone secretion, mitochondrial function, peroxisome biogenesis, translation elongation, stress response to cellular detoxification. Moreover, some yeast ABC proteins are orthologues of human disease genes, which makes yeast an excellent model system to study the molecular mechanisms of ABC protein-mediated disease. This review provides a comprehensive discussion and update on the function and transcriptional regulation of all known ABC genes from yeasts, including those discovered in fungal pathogens.

  19. ABC transporters in megakaryopoiesis and platelet activity.

    Science.gov (United States)

    Wang, Wei; Buitrago, Lorena; Wang, Ying

    2017-08-01

    ATP-binding cassette (ABC) is a family of transporters that facilitates the translocation of substrates across cell membrane using its ATPase subunit. These transporters have key roles in multidrug resistance, lipid homeostasis, antigen processing, immunity, cell proliferation and hematopoiesis. Some ABC transporters are selectively expressed on megakaryocyte progenitor, megakaryocyte and its cellular fragment platelet. However, the role of ABC transporters in hemostasis and thrombosis were not well explored until recently. Studies of both human genetic diseases and genetically-manipulated animal models have greatly improved our understanding of ABC transporters in regulating hematopoiesis particularly megakaryopoiesis and/or platelet activity. Human genome wide association studies (GWAS) have also unraveled the association between ABC transporters and thrombopoiesis in general population. Therefore, this review aims to summarize the recent advances in our understanding of how ABC transporters regulate megakaryopoiesis and platelet activity, the underlining mechanisms and their association with atherosclerosis and atherothrombosis. Last, the emerging therapeutic targets to slow down atherosclerosis development and prevent atherothrombosis via ABC transporters or downstream pathways will also be discussed. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Functional analysis of candidate ABC transporter proteins for sitosterol transport

    DEFF Research Database (Denmark)

    Albrecht, C; Elliott, J I; Sardini, A

    2002-01-01

    Two ATP-binding cassette (ABC) proteins, ABCG5 and ABCG8, have recently been associated with the accumulation of dietary cholesterol in the sterol storage disease sitosterolemia. These two 'half-transporters' are assumed to dimerize to form the complete sitosterol transporter which reduces...... the absorption of sitosterol and related molecules in the intestine by pumping them back into the lumen. Although mutations altering ABCG5 and ABCG8 are found in affected patients, no functional demonstration of sitosterol transport has been achieved. In this study, we investigated whether other ABC transporters......-specific ABC transporters have acquired specificity to exclude sitosterol and related sterols like cholesterol presumably because the abundance of cholesterol in the membrane would interfere with their action; in consequence, specific transporters have evolved to handle these sterols....

  1. Isolation and Characterization of the Colletotrichum acutatum ABC Transporter CaABC1

    Directory of Open Access Journals (Sweden)

    Suyoung Kim

    2014-12-01

    Full Text Available Fungi tolerate exposure to various abiotic stresses, including cytotoxic compounds and fungicides, via their ATP-driven efflux pumps belonging to ATP-binding cassette (ABC transporters. To clarify the molecular basis of interaction between the fungus and various abiotic stresses including fungicides, we constructed a cDNA library from germinated conidia of Colletotrichum acutatum, a major anthracnose pathogen of pepper (Capsicum annum L.. Over 1,000 cDNA clones were sequenced, of which single clone exhibited significant nucleotide sequence homology to ABC transporter genes. We isolated three fosmid clones containing the C. acutatum ABC1 (CaABC1 gene in full-length from genomic DNA library screening. The CaABC1 gene consists of 4,059 bp transcript, predicting a 1,353-aa protein. The gene contains the typical ABC signature and Walker A and B motifs. The 5′-flanking region contains a CAAT motif, a TATA box, and a Kozak region. Phylogenetic and structural analysis suggested that the CaABC1 is a typical ABC transporter gene highly conserved in various fungal species, as well as in Chromista, Metazoans, and Viridiplantae. We also found that CaABC1 was up-regulated during conidiation and a minimal medium condition. Moreover, CaABC1 was induced in iprobenfos, kresoxim-methyl, thiophanate-methyl, and hygromycin B. These results demonstrate that CaABC1 is necessary for conidiation, abiotic stress, and various fungicide resistances. These results will provide the basis for further study on the function of ABC transporter genes in C. acutatum.

  2. ABCdb: an ABC transporter database.

    Science.gov (United States)

    Quentin, Y; Fichant, G

    2000-10-01

    We present the first release of a database devoted to the ATP-binding cassette (ABC) protein domains (ABCdb). The ABC proteins are involved in a wide variety of physiological processes in Archea, Bacteria and Eucaryota where they are encoded by large families of paralogous genes. The majority of ABC domains energize the transport of compounds across the membranes. In bacteria, ABC transporters are involved in the uptake of a wide range of molecules and in mechanisms of virulence and antibiotic resistance. In eukaryotes, most of them are involved in drug resistance and in human cells, many are associated with diseases. Sequence analysis reveals that members of the ABC superfamily can be organized into sub-families and suggests that they have diverged from common ancestral forms. In this release, ABCdb includes the inventory and assembly of the ABC transporter systems of completely sequenced genomes. In addition to the protein entries, the database comprises information on functional domains, sequence motifs, predicted trans-membrane segments, and signal peptides. It also includes a classification in sub-families of the ABC systems as well as a classification of the different partners of the systems. Evolutionary trees and specific sequence patterns are provided for each sub-family. The database is endowed with a powerful query system and it was interfaced with blastP2 program for similarity searches. ABCdb has been developed in the ACeDB format, a database system developed by Jean Thierry-Mieg and Richard Durbin. ABCdb can be accessed via the World Wide Web (http://ir2lcb.cnrs-mrs.fr/ABCdb/).

  3. Backbone NMR resonance assignments of the nucleotide binding domain of the ABC multidrug transporter LmrA from Lactococcus lactis in its ADP-bound state.

    Science.gov (United States)

    Hellmich, Ute A; Duchardt-Ferner, Elke; Glaubitz, Clemens; Wöhnert, Jens

    2012-04-01

    LmrA from Lactococcus lactis is a multidrug transporter and a member of the ATP binding cassette (ABC) transporter family. ABC transporters consist of a transmembrane domain (TMD) and a nucleotide binding domain (NBD). The NBD contains the highly conserved signature motifs of this transporter superfamily. In the case of LmrA, the TMD and the NBD are expressed as a single polypeptide. LmrA catalyzes the extrusion of hydrophobic compounds including antibiotics from the cell membrane at the expense of ATP hydrolysis. ATP binds to the NBD, where binding and hydrolysis induce conformational changes that lead to the extrusion of the substrate via the TMD. Here, we report the (1)H, (13)C and (15)N backbone chemical shift assignments of the isolated 263 amino acid containing NBD of LmrA in its ADP bound state.

  4. Yeast ABC transporters in lipid trafficking.

    Science.gov (United States)

    Prasad, Rajendra; Khandelwal, Nitesh Kumar; Banerjee, Atanu

    2016-08-01

    Throughout its evolution, the ATP-binding cassette (ABC) transporter superfamily has experienced a rapid expansion in its substrate repertoire and functions. Of the diverse functions that these pumps offer, their drug transport properties have attracted considerable attention primarily owing to their clinical significance. Despite this fact, emerging evidence suggests that physiological substrates of transporters also affect the overall functioning of an organism. Lipids, as substrates of ABC transporters, constitute one feature found in all representative groups of the living kingdom. Due to the importance of lipid species in the cellular physiology of an organism, their proper distribution within cells is crucial. This fact is well exemplified by the vast number of medical conditions that have been caused as a result of perturbations in ABC transporter-mediated lipid transport in higher organisms. In yeasts, apart from providing transport functions, ABC transporters also coordinate regulatory networks with lipids. This review focuses on yeast ABC transporters involved in the transport of lipids and briefly discusses the integration of their regulatory network with that of the lipid species. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. The Schizosaccharomyces pombe mam1 gene encodes an ABC transporter mediating secretion of M-factor

    DEFF Research Database (Denmark)

    Christensen, P U; Davey, William John; Nielsen, O

    1997-01-01

    In the fission yeast Schizosaccharomyces pombe, cells of opposite mating type communicate via diffusible peptide pheromones prior to mating. We have cloned the S. pombe mam1 gene, which encodes a 1336-amino acid protein belonging to the ATP-binding cassette (ABC) superfamily. The mam1 gene is only...... expressed in M cells and the gene product is responsible for the secretion of the mating pheromone. M-factor, a nonapeptide that is S-farnesylated and carboxy-methylated on its C-terminal cysteine residue. The predicted Mam1 protein is highly homologous to mammalian multiple drug-resistance proteins...

  6. Effect of Oxidative Stress on ABC Transporters: Contribution to Epilepsy Pharmacoresistance

    Directory of Open Access Journals (Sweden)

    Gurpreet Kaur Grewal

    2017-02-01

    Full Text Available Epilepsy is a neurological disorder affecting around 1%–2% of population worldwide and its treatment includes use of antiepileptic drugs to control seizures. Failure to respond to antiepileptic drug therapy is a major clinical problem and over expression of ATP-binding cassette transporters is considered one of the major reasons for pharmacoresistance. In this review, we have summarized the regulation of ABC transporters in response to oxidative stress due to disease and antiepileptic drugs. Further, ketogenic diet and antioxidants were examined for their role in pharmacoresistance. The understanding of signalling pathways and mechanism involved may help in identifying potential therapeutic targets and improving drug response.

  7. Purification and biochemical characterisation of the yeast ABC transporter Pdr11p

    DEFF Research Database (Denmark)

    Laub, Katrine Rude

    Sterols constitute an essential lipid class in eukaryotic membranes where intracellular distributions are highly regulated. In the yeast Saccharomyces cerevisiae sterol uptake has been attributed to the two plasma membrane-localised ATP-binding cassette (ABC) transporters, Aus1p and Pdr11p...... understanding of their action and molecular functioning, it is helpful to focus on homogeneous preparations of membrane proteins reconstituted into model membranes like detergent micelles, liposomes, and nanodiscs. In the present study, I report the first successful expression, purification, and reconstitution...

  8. ABC Transporters and the Alzheimer’s Disease Enigma

    Directory of Open Access Journals (Sweden)

    Andrea eWolf

    2012-06-01

    Full Text Available Alzheimer’s disease (AD is considered the disease of the 21st century. With a 10-fold increase in global incidence over the past 100 years, AD is now reaching epidemic proportions and by all projections, AD patient numbers will continue to rise. Despite intense research efforts, AD remains a mystery and effective therapies are still unavailable. This represents an unmet need resulting in clinical, social, and economic problems.Over the last decade, a new AD research focus has emerged: ATP-binding cassette (ABC transporters. In this article, we provide an overview of the ABC transporters ABCA1, ABCA2, P-glycoprotein (ABCB1, Mrp1 (ABCC1 and BCRP (ABCG2, all of which are expressed in the brain and have been implicated in AD. We summarize recent findings on the role of these five transporters in AD, and discuss their potential to serve as therapeutic targets.

  9. ABC transporters as multidrug resistance mechanisms and the development of chemosensitizers for their reversal

    Directory of Open Access Journals (Sweden)

    Choi Cheol-Hee

    2005-10-01

    Full Text Available Abstract One of the major problems related with anticancer chemotherapy is resistance against anticancer drugs. The ATP-binding cassette (ABC transporters are a family of transporter proteins that are responsible for drug resistance and a low bioavailability of drugs by pumping a variety of drugs out cells at the expense of ATP hydrolysis. One strategy for reversal of the resistance of tumor cells expressing ABC transporters is combined use of anticancer drugs with chemosensitizers. In this review, the physiological functions and structures of ABC transporters, and the development of chemosensitizers are described focusing on well-known proteins including P-glycoprotein, multidrug resistance associated protein, and breast cancer resistance protein.

  10. The mitochondrial transporter ABC-me (ABCB10), a downstream target of GATA-1, is essential for erythropoiesis in vivo

    Science.gov (United States)

    Hyde, B B; Liesa, M; Elorza, A A; Qiu, W; Haigh, S E; Richey, L; Mikkola, H K; Schlaeger, T M; Shirihai, O S

    2012-01-01

    The mitochondrial transporter ATP binding cassette mitochondrial erythroid (ABC-me/ABCB10) is highly induced during erythroid differentiation by GATA-1 and its overexpression increases hemoglobin production rates in vitro. However, the role of ABC-me in erythropoiesis in vivo is unknown. Here we report for the first time that erythrocyte development in mice requires ABC-me. ABC-me−/− mice die at day 12.5 of gestation, showing nearly complete eradication of primitive erythropoiesis and lack of hemoglobinized cells at day 10.5. ABC-me−/− erythroid cells fail to differentiate because they exhibit a marked increase in apoptosis, both in vivo and ex vivo. Erythroid precursors are particularly sensitive to oxidative stress and ABC-me in the heart and its yeast ortholog multidrug resistance-like 1 have been shown to protect against oxidative stress. Thus, we hypothesized that increased apoptosis in ABC-me−/− erythroid precursors was caused by oxidative stress. Within this context, ABC-me deletion causes an increase in mitochondrial superoxide production and protein carbonylation in erythroid precursors. Furthermore, treatment of ABC-me−/− erythroid progenitors with the mitochondrial antioxidant MnTBAP (superoxide dismutase 2 mimetic) supports survival, ex vivo differentiation and increased hemoglobin production. Altogether, our findings demonstrate that ABC-me is essential for erythropoiesis in vivo. PMID:22240895

  11. Importance of ABC Transporters in Drug Development.

    Science.gov (United States)

    Benadiba, Marcel; Maor, Yehoshua

    2016-01-01

    ATP-binding cassette (ABC) transporters are a huge family of ATP-dependent transmembrane proteins whose main function is exporting or importing substances or molecules through the cell membranes, plasma cell membrane, or inner membranes in organelles. They fulfill these functions by maintaining cell integrity, metabolism, and homeostasis. They are expressed in a variety of tissues as they transport numerous essential compounds including lipids and other signaling molecules. ABC transporters became widely studied since the discovery of their ability to carry a multitude of xenobiotics, including therapeutic drugs, and in light of the fact that they represent a hurdle for the treatment of resistant cancers. In contrast, the role of ABC transporters in neurological diseases like Alzheimer`s and Parkinson`s, depression, schizophrenia, and epilepsy remains controversial and their mechanism of action in these pathologies remains elusive, thus hindering the implementation of therapies aimed at modulating the functions of these transporters. To date, a number of natural and synthetic compounds are known to act as inhibitors, substrates, and even inducers of these transporters, being able to modulate their expression and/or function; however, their implication as therapeutic agents is far from reaching wide clinical utilization. This review highlights the importance of overcoming the challenges posed by ABC transporters in drug development.

  12. ABC Transport Proteins in Cardiovascular Disease-A Brief Summary.

    Science.gov (United States)

    Schumacher, Toni; Benndorf, Ralf A

    2017-04-06

    Adenosine triphosphate (ATP)-binding cassette (ABC) transporters may play an important role in the pathogenesis of atherosclerotic vascular diseases due to their involvement in cholesterol homeostasis, blood pressure regulation, endothelial function, vascular inflammation, as well as platelet production and aggregation. In this regard, ABC transporters, such as ABCA1, ABCG5 and ABCG8, were initially found to be responsible for genetically-inherited syndromes like Tangier diseases and sitosterolemia. These findings led to the understanding of those transporter's function in cellular cholesterol efflux and thereby also linked them to atherosclerosis and cardiovascular diseases (CVD). Subsequently, further ABC transporters, i.e., ABCG1, ABCG4, ABCB6, ABCC1, ABCC6 or ABCC9, have been shown to directly or indirectly affect cellular cholesterol efflux, the inflammatory response in macrophages, megakaryocyte proliferation and thrombus formation, as well as vascular function and blood pressure, and may thereby contribute to the pathogenesis of CVD and its complications. Furthermore, ABC transporters, such as ABCB1, ABCC2 or ABCG2, may affect the safety and efficacy of several drug classes currently in use for CVD treatment. This review will give a brief overview of ABC transporters involved in the process of atherogenesis and CVD pathology. It also aims to briefly summarize the role of ABC transporters in the pharmacokinetics and disposition of drugs frequently used to treat CVD and CVD-related complications.

  13. Structural basis for the mechanism of ABC transporters.

    Science.gov (United States)

    Beis, Konstantinos

    2015-10-01

    The ATP-binding cassette (ABC) transporters are primary transporters that couple the energy stored in adenosine triphosphate (ATP) to the movement of molecules across the membrane. ABC transporters can be divided into exporters and importers; importers mediate the uptake of essential nutrients into cells and are found predominantly in prokaryotes whereas exporters transport molecules out of cells or into organelles and are found in all organisms. ABC exporters have been linked with multi-drug resistance in both bacterial and eukaryotic cells. ABC transporters are powered by the hydrolysis of ATP and transport their substrate via the alternating access mechanism, whereby the protein alternates between a conformation in which the substrate-binding site is accessible from the outside of the membrane, outward-facing and one in which it is inward-facing. In this mini-review, the structures of different ABC transporter types in different conformations are presented within the context of the alternating access mechanism and how they have shaped our current understanding of the mechanism of ABC transporters. © 2015 Authors; published by Portland Press Limited.

  14. ABC Transporters in Cancer Stem Cells: Beyond Chemoresistance.

    Science.gov (United States)

    Begicevic, Romana-Rea; Falasca, Marco

    2017-11-08

    The efficacy of chemotherapy is one of the main challenges in cancer treatment and one of the major obstacles to overcome in achieving lasting remission and a definitive cure in patients with cancer is the emergence of cancer resistance. Indeed, drug resistance is ultimately accountable for poor treatment outcomes and tumour relapse. There are various molecular mechanisms involved in multidrug resistance, such as the change in the activity of membrane transporters primarily belonging to the ATP binding cassette (ABC) transporter family. In addition, it has been proposed that this common feature could be attributed to a subpopulation of slow-cycling cancer stem cells (CSCs), endowed with enhanced tumorigenic potential and multidrug resistance. CSCs are characterized by the overexpression of specific surface markers that vary in different cancer cell types. Overexpression of ABC transporters has been reported in several cancers and more predominantly in CSCs. While the major focus on the role played by ABC transporters in cancer is polarized by their involvement in chemoresistance, emerging evidence supports a more active role of these proteins, in which they release specific bioactive molecules in the extracellular milieu. This review will outline our current understanding of the role played by ABC transporters in CSCs, how their expression is regulated and how they support the malignant metabolic phenotype. To summarize, we suggest that the increased expression of ABC transporters in CSCs may have precise functional roles and provide the opportunity to target, particularly these cells, by using specific ABC transporter inhibitors.

  15. ABCE1 is a highly conserved RNA silencing suppressor.

    Directory of Open Access Journals (Sweden)

    Kairi Kärblane

    Full Text Available ATP-binding cassette sub-family E member 1 (ABCE1 is a highly conserved protein among eukaryotes and archaea. Recent studies have identified ABCE1 as a ribosome-recycling factor important for translation termination in mammalian cells, yeast and also archaea. Here we report another conserved function of ABCE1. We have previously described AtRLI2, the homolog of ABCE1 in the plant Arabidopsis thaliana, as an endogenous suppressor of RNA silencing. In this study we show that this function is conserved: human ABCE1 is able to suppress RNA silencing in Nicotiana benthamiana plants, in mammalian HEK293 cells and in the worm Caenorhabditis elegans. Using co-immunoprecipitation and mass spectrometry, we found a number of potential ABCE1-interacting proteins that might support its function as an endogenous suppressor of RNA interference. The interactor candidates are associated with epigenetic regulation, transcription, RNA processing and mRNA surveillance. In addition, one of the identified proteins is translin, which together with its binding partner TRAX supports RNA interference.

  16. ABC Transporters in Cancer Stem Cells: Beyond Chemoresistance

    Directory of Open Access Journals (Sweden)

    Romana-Rea Begicevic

    2017-11-01

    Full Text Available The efficacy of chemotherapy is one of the main challenges in cancer treatment and one of the major obstacles to overcome in achieving lasting remission and a definitive cure in patients with cancer is the emergence of cancer resistance. Indeed, drug resistance is ultimately accountable for poor treatment outcomes and tumour relapse. There are various molecular mechanisms involved in multidrug resistance, such as the change in the activity of membrane transporters primarily belonging to the ATP binding cassette (ABC transporter family. In addition, it has been proposed that this common feature could be attributed to a subpopulation of slow-cycling cancer stem cells (CSCs, endowed with enhanced tumorigenic potential and multidrug resistance. CSCs are characterized by the overexpression of specific surface markers that vary in different cancer cell types. Overexpression of ABC transporters has been reported in several cancers and more predominantly in CSCs. While the major focus on the role played by ABC transporters in cancer is polarized by their involvement in chemoresistance, emerging evidence supports a more active role of these proteins, in which they release specific bioactive molecules in the extracellular milieu. This review will outline our current understanding of the role played by ABC transporters in CSCs, how their expression is regulated and how they support the malignant metabolic phenotype. To summarize, we suggest that the increased expression of ABC transporters in CSCs may have precise functional roles and provide the opportunity to target, particularly these cells, by using specific ABC transporter inhibitors.

  17. Role of ATP binding and hydrolysis in assembly of MacAB-TolC macrolide transporter

    Science.gov (United States)

    Lu, Shuo; Zgurskaya, Helen I.

    2012-01-01

    Summary MacB is a founding member of the Macrolide Exporter family of transporters belonging to the ATP-Binding Cassette superfamily. These proteins are broadly represented in genomes of both gram-positive and gram-negative bacteria and are implicated in virulence and protection against antibiotics and peptide toxins. MacB transporter functions together with MacA, a periplasmic membrane fusion protein, which stimulates MacB ATPase. In gram-negative bacteria, MacA is believed to couple ATP hydrolysis to transport of substrates across the outer membrane through a TolC-like channel. In this study, we report a real-time analysis of concurrent ATP hydrolysis and assembly of MacAB-TolC complex. MacB binds nucleotides with a low millimolar affinity and fast on- and off-rates. In contrast, MacA-MacB complex is formed with a nanomolar affinity, which further increases in the presence of ATP. Our results strongly suggest that association between MacA and MacB is stimulated by ATP binding to MacB but remains unchanged during ATP hydrolysis cycle. We also found that the large periplasmic loop of MacB plays the major role in coupling reactions separated in two different membranes. This loop is required for MacA-dependent stimulation of MacB ATPase and at the same time, contributes to recruitment of TolC into a trans-envelope complex. PMID:23057817

  18. Current advances on ABC drug transporters in fish.

    Science.gov (United States)

    Luckenbach, Till; Fischer, Stephan; Sturm, Armin

    2014-09-01

    Most members of the large ATP-binding cassette (ABC) gene family are transporters involved in substrate translocation across biological membranes. In eukaryotes, ABC proteins functioning as drug transporters are located in the plasma membrane and mediate the cellular efflux of a wide range of organic chemicals, with some transporters also transporting certain metals. As the enhanced expression of ABC drug transporters can confer multidrug resistance (MDR) to cancers and multixenobiotic resistance (MXR) to organisms from polluted habitats, these ABC family members are also referred to as MDR or MXR proteins. In mammals, ABC drug transporters show predominant expression in tissues involved in excretion or constituting internal or external body boundaries, where they facilitate the excretion of chemicals and their metabolites, and limit chemical uptake and penetration into "sanctuary" sites of the body. Available knowledge about ABC proteins is still limited in teleost fish, a large vertebrate group of high ecological and economic importance. Using transport activity measurements and immunochemical approaches, early studies demonstrated similarities in the tissue distribution of ABC drug transporters between teleosts and mammals, suggesting conserved roles of the transporters in the biochemical defence against toxicants. Recently, the availability of teleost genome assemblies has stimulated studies of the ABC family in this taxon. This review summarises the current knowledge regarding the genetics, functional properties, physiological function, and ecotoxicological relevance of teleostean ABC transporters. The available literature is reviewed with emphasis on recent studies addressing the tissue distribution, substrate spectrum, regulation, physiological function and phylogenetic origin of teleostean ABC transporters. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  19. Inventory, assembly and analysis of Bacillus subtilis ABC transport systems.

    Science.gov (United States)

    Quentin, Y; Fichant, G; Denizot, F

    1999-04-02

    We have undertaken the inventory and assembly of the ATP binding cassette (ABC) transporter systems in the complete genome of Bacillus subtilis. We combined the identification of the three protein partners that compose an ABC transporter (nucleotide-binding domain, NBD; membrane spanning domain, MSD; and solute-binding protein, SBP) with constraints on the genetic organization. This strategy allowed the identification of 86 NBDs in 78 proteins, 103 MSD proteins and 37 SBPs. The analysis of transcriptional units allows the reconstruction of 59 ABC transporters, which include at least one NBD and one MSD. A particular class of five dimeric ATPases was not associated to MSD partners and is assumed to be involved either in macrolide resistance or regulation of translation elongation. In addition, we have detected five genes encoding ATPases without any gene coding for MSD protein in their neighborhood and 11 operons that encode only the membrane and solute-binding proteins. On the bases of similarities, three ATP-binding proteins are proposed to energize ten incomplete systems, suggesting that one ATPase may be recruited by more than one transporter. Finally, we estimate that the B. subtilis genome encodes for at least 78 ABC transporters that have been split in 38 importers and 40 extruders. The ABC systems have been further classified into 11 sub-families according to the tree obtained from the NBDs and the clustering of the MSDs and the SBPs. Comparisons with Escherichia coli show that the extruders are over-represented in B. subtilis, corresponding to an expansion of the sub-families of antibiotic and drug resistance systems. Copyright 1999 Academic Press.

  20. Supplementary data: Novel mutation in ATP-binding domain of ...

    Indian Academy of Sciences (India)

    Novel mutation in ATP-binding domain of ABCD1 gene in adrenoleucodystrophy. Neeraj Kumar, Krishna K. Taneja, Atul Kumar, Deepti Nayar, Bhupesh Taneja, Satindra Aneja,. Madhuri Behari, Veena Kalra and Surendra K. Bansal. J. Genet. 89, 473–477. Figure 1. Rmsd plot of native and Arg617Ser substituted models.

  1. Computer-Aided Recognition of ABC Transporters Substrates and Its Application to the Development of New Drugs for Refractory Epilepsy.

    Science.gov (United States)

    Couyoupetrou, Manuel; Gantner, Melisa E; Di Ianni, Mauricio E; Palestro, Pablo H; Enrique, Andrea V; Gavernet, Luciana; Ruiz, Maria E; Pesce, Guido; Bruno-Blanch, Luis E; Talevi, Alan

    2017-01-01

    Despite the introduction of more than 15 third generation antiepileptic drugs to the market from 1990 to the moment, about one third of the epileptic patients still suffer from refractory to intractable epilepsy. Several hypotheses seek to explain the failure of drug treatments to control epilepsy symptoms in such patients. The most studied one proposes that drug resistance might be related with regional overactivity of efflux transporters from the ATP-Binding Cassette (ABC) superfamily at the blood-brain barrier and/or the epileptic foci in the brain. Different strategies have been conceived to address the transporter hypothesis, among them inhibiting or down-regulating the efflux transporters or bypassing them through a diversity of artifices. Here, we review scientific evidence supporting the transporter hypothesis along with its limitations, as well as computer-assisted early recognition of ABC transporter substrates as an interesting strategy to develop novel antiepileptic drugs capable of treating refractory epilepsy linked to ABC transporters overactivity.

  2. Genome-wide comparative analysis of ABC systems in the Bdellovibrio-and-like organisms.

    Science.gov (United States)

    Li, Nan; Chen, Huan; Williams, Henry N

    2015-05-10

    Bdellovibrio-and-like organisms (BALOs) are gram-negative, predatory bacteria with wide variations in genome sizes and GC content and ecological habitats. The ATP-binding cassette (ABC) systems have been identified in several prokaryotes, fungi and plants and have a role in transport of materials in and out of cells and in cellular processes. However, knowledge of the ABC systems of BALOs remains obscure. A total of 269 putative ABC proteins were identified in BALOs. The genes encoding these ABC systems occupy nearly 1.3% of the gene content in freshwater Bdellovibrio strains and about 0.7% in their saltwater counterparts. The proteins found belong to 25 ABC system families based on their structural characteristics and functions. Among these, 16 families function as importers, 6 as exporters and 3 are involved in various cellular processes. Eight of these 25 ABC system families were deduced to be the core set of ABC systems conserved in all BALOs. All Bacteriovorax strains have 28 or less ABC systems. On the contrary, the freshwater Bdellovibrio strains have more ABC systems, typically around 51. In the genome of Bdellovibrio exovorus JSS (CP003537.1), 53 putative ABC systems were detected, representing the highest number among all the BALO genomes examined in this study. Unexpected high numbers of ABC systems involved in cellular processes were found in all BALOs. Phylogenetic analysis suggests that the majority of ABC proteins can be assigned into many separate families with high bootstrap supports (>50%). In this study, a general framework of sequence-structure-function connections for the ABC systems in BALOs was revealed providing novel insights for future investigations. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. Comparison of mechanistic transport cycle models of ABC exporters.

    Science.gov (United States)

    Szöllősi, Dániel; Rose-Sperling, Dania; Hellmich, Ute A; Stockner, Thomas

    2018-04-01

    ABC (ATP binding cassette) transporters, ubiquitous in all kingdoms of life, carry out essential substrate transport reactions across cell membranes. Their transmembrane domains bind and translocate substrates and are connected to a pair of nucleotide binding domains, which bind and hydrolyze ATP to energize import or export of substrates. Over four decades of investigations into ABC transporters have revealed numerous details from atomic-level structural insights to their functional and physiological roles. Despite all these advances, a comprehensive understanding of the mechanistic principles of ABC transporter function remains elusive. The human multidrug resistance transporter ABCB1, also referred to as P-glycoprotein (P-gp), is one of the most intensively studied ABC exporters. Using ABCB1 as the reference point, we aim to compare the dominating mechanistic models of substrate transport and ATP hydrolysis for ABC exporters and to highlight the experimental and computational evidence in their support. In particular, we point out in silico studies that enhance and complement available biochemical data. "This article is part of a Special Issue entitled: Beyond the Structure-Function Horizon of Membrane Proteins edited by Ute Hellmich, Rupak Doshi and Benjamin McIlwain." Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  4. The Arabidopsis thaliana ABC protein superfamily, a complete inventory.

    Science.gov (United States)

    Sánchez-Fernández, R; Davies, T G; Coleman, J O; Rea, P A

    2001-08-10

    We describe the first complete inventory of ATP-binding cassette (ABC) proteins from a multicellular organism, the model plant Arabidopsis thaliana. By the application of several search criteria, Arabidopsis was found to contain a total of 129 open reading frames (ORFs) capable of encoding ABC proteins, of which 103 possessed contiguous transmembrane spans and were identified as putative intrinsic membrane proteins. Fifty-two of the putative intrinsic membrane proteins contained at least two transmembrane domains (TMDs) and two nucleotide-binding folds (NBFs) and could be classified as belonging to one of five subfamilies of full-molecule transporters. The other 51 putative membrane proteins, all of which were half-molecule transporters, fell into five subfamilies. Of the remaining ORFs identified, all of which encoded proteins lacking TMDs, 11 could be classified into three subfamilies. There were no obvious homologs in other organisms for 15 of the ORFs which encoded a heterogeneous group of non-intrinsic ABC proteins (NAPs). Unrooted phylogenetic analyses substantiated the subfamily designations. Notable features of the Arabidopsis ABC superfamily was the presence of a large yeast-like PDR subfamily, and the absence of genes encoding bona fide cystic fibrosis transmembrane conductance regulator (CFTR), sulfonylurea receptor (SUR), and heavy metal tolerance factor 1 (HMT1) homologs. Arabidopsis was unusual in its large allocation of ORFs (a minimum of 0.5%) to members of the ABC protein superfamily.

  5. The AbcA Transporter of Staphylococcus aureus Affects Cell Autolysis

    Science.gov (United States)

    Schrader-Fischer, Gesine; Berger-Bächi, Brigitte

    2001-01-01

    Increased production of penicillin-binding protein PBP 4 is known to increase peptidoglycan cross-linking and contributes to methicillin resistance in Staphylococcus aureus. The pbp4 gene shares a 400-nucleotide intercistronic region with the divergently transcribed abcA gene, encoding an ATP-binding cassette transporter of unknown function. Our study revealed that methicillin stimulated abcA transcription but had no effects on pbp4 transcription. Analysis of abcA expression in mutants defective for global regulators showed that abcA is under the control of agr. Insertional inactivation of abcA by an erythromycin resistance determinant did not influence pbp4 transcription, nor did it alter resistance to methicillin and other cell wall-directed antibiotics. However, abcA mutants showed spontaneous partial lysis on plates containing subinhibitory concentrations of methicillin due to increased spontaneous autolysis. Since the autolytic zymograms of cell extracts were identical in mutants and parental strains, we postulate an indirect role of AbcA in control of autolytic activities and in protection of the cells against methicillin. PMID:11158733

  6. Brain barriers and functional interfaces with sequential appearance of ABC efflux transporters during human development

    DEFF Research Database (Denmark)

    Møllgård, Kjeld; Dziegielewska, Katarzyna M.; Holst, Camilla B.

    2017-01-01

    Adult brain is protected from entry of drugs and toxins by specific mechanisms such as ABC (ATP-binding Cassette) efflux transporters. Little is known when these appear in human brain during development. Cellular distribution of three main ABC transporters (ABCC1, ABCG2, ABCB1) was determined...... at blood-brain barriers and interfaces in human embryos and fetuses in first half of gestation. Antibodies against claudin-5 and-11 and antibodies to α-fetoprotein were used to describe morphological and functional aspects of brain barriers. First exchange interfaces to be established, probably at 4......-5 weeks post conception, are between brain and embryonic cerebrospinal fluid (eCSF) and between outer surface of brain anlage and primary meninx. They already exclude α-fetoprotein and are immunopositive for both claudins, ABCC1 and ABCG2. ABCB1 is detectable within a week of blood vessels first...

  7. Obstacles to Brain Tumor Therapy: Key ABC Transporters

    Directory of Open Access Journals (Sweden)

    Juwina Wijaya

    2017-11-01

    Full Text Available The delivery of cancer chemotherapy to treat brain tumors remains a challenge, in part, because of the inherent biological barrier, the blood–brain barrier. While its presence and role as a protector of the normal brain parenchyma has been acknowledged for decades, it is only recently that the important transporter components, expressed in the tightly knit capillary endothelial cells, have been deciphered. These transporters are ATP-binding cassette (ABC transporters and, so far, the major clinically important ones that functionally contribute to the blood–brain barrier are ABCG2 and ABCB1. A further limitation to cancer therapy of brain tumors or brain metastases is the blood–tumor barrier, where tumors erect a barrier of transporters that further impede drug entry. The expression and regulation of these two transporters at these barriers, as well as tumor derived alteration in expression and/or mutation, are likely obstacles to effective therapy.

  8. Polymorphism in ABC transporter genes of Dirofilaria immitis

    Directory of Open Access Journals (Sweden)

    Thangadurai Mani

    2017-08-01

    Full Text Available Dirofilaria immitis, a filarial nematode, causes dirofilariasis in dogs, cats and occasionally in humans. Prevention of the disease has been mainly by monthly use of the macrocyclic lactone (ML endectocides during the mosquito transmission season. Recently, ML resistance has been confirmed in D. immitis and therefore, there is a need to find new classes of anthelmintics. One of the mechanisms associated with ML resistance in nematodes has been the possible role of ATP binding cassette (ABC transporters in reducing drug concentrations at receptor sites. ABC transporters, mainly from sub-families B, C and G, may contribute to multidrug resistance (MDR by active efflux of drugs out of the cell. Gene products of ABC transporters may thus serve as the targets for agents that may modulate susceptibility to drugs, by inhibiting drug transport. ABC transporters are believed to be involved in a variety of physiological functions critical to the parasite, such as sterol transport, and therefore may also serve as the target for drugs that can act as anthelmintics on their own. Knowledge of polymorphism in these ABC transporter genes in nematode parasites could provide useful information for the process of drug design. We have identified 15 ABC transporter genes from sub-families A, B, C and G, in D. immitis, by comparative genomic approaches and analyzed them for polymorphism. Whole genome sequencing data from four ML susceptible (SUS and four loss of efficacy (LOE pooled populations were used for single nucleotide polymorphism (SNP genotyping. Out of 231 SNPs identified in those 15 ABC transporter genes, 89 and 75 of them were specific to the SUS or LOE populations, respectively. A few of the SNPs identified may affect gene expression, protein function, substrate specificity or resistance development and may be useful for transporter inhibitor/anthelmintic drug design, or in order to anticipate resistance development. Keywords: Dirofilaria immitis

  9. Export of recombinant proteins in Escherichia coli using ABC transporter with an attached lipase ABC transporter recognition domain (LARD

    Directory of Open Access Journals (Sweden)

    Moon Yuseok

    2009-01-01

    Full Text Available Abstract Background ATP binding cassette (ABC transporter secretes the protein through inner and outer membranes simultaneously in gram negative bacteria. Thermostable lipase (TliA of Pseudomonas fluorescens SIK W1 is secreted through the ABC transporter. TliA has four glycine-rich repeats (GGXGXD in its C-terminus, which appear in many ABC transporter-secreted proteins. From a homology model of TliA derived from the structure of P. aeruginosa alkaline protease (AprA, lipase ABC transporter domains (LARDs were designed for the secretion of fusion proteins. Results The LARDs included four glycine-rich repeats comprising a β-roll structure, and were added to the C-terminus of test proteins. Either Pro-Gly linker or Factor Xa site was added between fusion proteins and LARDs. We attached different length of LARDs such as LARD0, LARD1 or whole TliA (the longest LARD to three types of proteins; green fluorescent protein (GFP, epidermal growth factor (EGF and cytoplasmic transduction peptide (CTP. These fusion proteins were expressed in Escherichia coli together with ABC transporter of either P. fluorescens or Erwinia chrysanthemi. Export of fusion proteins with the whole TliA through the ABC transporter was evident on the basis of lipase enzymatic activity. Upon supplementation of E. coli with ABC transporter, GFP-LARDs and EGF-LARDs were excreted into the culture supernatant. Conclusion The LARDs or whole TliA were attached to C-termini of model proteins and enabled the export of the model proteins such as GFP and EGF in E. coli supplemented with ABC transporter. These results open the possibility for the extracellular production of recombinant proteins in Pseudomonas using LARDs or TliA as a C-terminal signal sequence.

  10. ABC transporters in fish species: a review

    Directory of Open Access Journals (Sweden)

    Marta eFerreira

    2014-07-01

    Full Text Available ATP-binding cassette (ABC proteins were first recognized for their role in multidrug resistance (MDR in chemotherapeutic treatments, which is a major impediment for the successful treatment of many forms of malignant tumors in humans. These proteins, highly conserved throughout vertebrate species, were later related to cellular detoxification and accounted as responsible for protecting aquatic organisms from xenobiotic insults in the so-called multixenobiotic resistance mechanism (MXR. In recent years, research on these proteins in aquatic species has highlighted their importance in the detoxification mechanisms in fish thus it is of extreme added value to continue these studies. Several transporters have been pointed out as relevant in the ecotoxicological context associated to the transport of xenobiotics, such as P-glycoproteins (Pgps, multidrug-resistance-associated proteins (MRPs 1-5 and breast resistance associated protein (BCRP. In mammals, several nuclear receptors have been identified as mediators of phase I and II metabolizing enzymes and ABC transporters. In aquatic species, knowledge on co-regulation of detoxification mechanism is scarce and needs to be addressed. The interaction of emergent contaminants, with chemosensitizer potential, with ABC transporters in aquatic organisms can compromise detoxification processes and have population effects and should be studied in more detail. This review intends to summarize the recent advances in research on MXR mechanisms in fish species, focusing in 1 regulation and functioning of ABC proteins; 2 cooperation with phase I and II biotransformation enzymes; and 3 ecotoxicological relevance and information on emergent pollutants with ability to modulate ABC transporters expression and activity. Several lines of evidence are clear suggesting the important role of these transporters in detoxification mechanisms and must be further investigated in fish.

  11. A burst of ABC genes in the genome of the polyphagous spider mite Tetranychus urticae

    Science.gov (United States)

    2013-01-01

    Background The ABC (ATP-binding cassette) gene superfamily is widespread across all living species. The majority of ABC genes encode ABC transporters, which are membrane-spanning proteins capable of transferring substrates across biological membranes by hydrolyzing ATP. Although ABC transporters have often been associated with resistance to drugs and toxic compounds, within the Arthropoda ABC gene families have only been characterized in detail in several insects and a crustacean. In this study, we report a genome-wide survey and expression analysis of the ABC gene superfamily in the spider mite, Tetranychus urticae, a chelicerate ~ 450 million years diverged from other Arthropod lineages. T. urticae is a major agricultural pest, and is among of the most polyphagous arthropod herbivores known. The species resists a staggering array of toxic plant secondary metabolites, and has developed resistance to all major classes of pesticides in use for its control. Results We identified 103 ABC genes in the T. urticae genome, the highest number discovered in a metazoan species to date. Within the T. urticae ABC gene set, all members of the eight currently described subfamilies (A to H) were detected. A phylogenetic analysis revealed that the high number of ABC genes in T. urticae is due primarily to lineage-specific expansions of ABC genes within the ABCC, ABCG and ABCH subfamilies. In particular, the ABCC subfamily harbors the highest number of T. urticae ABC genes (39). In a comparative genomic analysis, we found clear orthologous relationships between a subset of T. urticae ABC proteins and ABC proteins in both vertebrates and invertebrates known to be involved in fundamental cellular processes. These included members of the ABCB-half transporters, and the ABCD, ABCE and ABCF families. Furthermore, one-to-one orthologues could be distinguished between T. urticae proteins and human ABCC10, ABCG5 and ABCG8, the Drosophila melanogaster sulfonylurea receptor and ecdysone

  12. Polymorphism in ABC transporter genes of Dirofilaria immitis.

    Science.gov (United States)

    Mani, Thangadurai; Bourguinat, Catherine; Prichard, Roger K

    2017-08-01

    Dirofilaria immitis, a filarial nematode, causes dirofilariasis in dogs, cats and occasionally in humans. Prevention of the disease has been mainly by monthly use of the macrocyclic lactone (ML) endectocides during the mosquito transmission season. Recently, ML resistance has been confirmed in D. immitis and therefore, there is a need to find new classes of anthelmintics. One of the mechanisms associated with ML resistance in nematodes has been the possible role of ATP binding cassette (ABC) transporters in reducing drug concentrations at receptor sites. ABC transporters, mainly from sub-families B, C and G, may contribute to multidrug resistance (MDR) by active efflux of drugs out of the cell. Gene products of ABC transporters may thus serve as the targets for agents that may modulate susceptibility to drugs, by inhibiting drug transport. ABC transporters are believed to be involved in a variety of physiological functions critical to the parasite, such as sterol transport, and therefore may also serve as the target for drugs that can act as anthelmintics on their own. Knowledge of polymorphism in these ABC transporter genes in nematode parasites could provide useful information for the process of drug design. We have identified 15 ABC transporter genes from sub-families A, B, C and G, in D. immitis, by comparative genomic approaches and analyzed them for polymorphism. Whole genome sequencing data from four ML susceptible (SUS) and four loss of efficacy (LOE) pooled populations were used for single nucleotide polymorphism (SNP) genotyping. Out of 231 SNPs identified in those 15 ABC transporter genes, 89 and 75 of them were specific to the SUS or LOE populations, respectively. A few of the SNPs identified may affect gene expression, protein function, substrate specificity or resistance development and may be useful for transporter inhibitor/anthelmintic drug design, or in order to anticipate resistance development. Copyright © 2017 The Authors. Published by

  13. Polymorphism in a second ABC transproter gene located within the class II region of the human major histocompatibility complex

    Energy Technology Data Exchange (ETDEWEB)

    Powis, S.H.; Mockridge, I.; Kelly, A.; Glynne, R.; Beck, S.; Trowsdale, J. (Imperial Cancer Research Fund Labs., London (United Kingdom)); Kerr, L.A. (Guy' s Campus, London (United Kingdom)); Gileadi, U. (Univ. of Oxford (United Kingdom))

    1992-02-15

    Recent studies have identified genes within the major histocompatibility complex (MHC) that may play a role in presentation of antigenic peptides to T cells. The authors have previously described RING4, a gene within the human MHC class II region that has sequence homology with members of the ABC (ATP-binding cassette) transporter superfamily. They now report the nucleotide sequence of RING11, a second ABC transporter gene located approximately 7 kilobases telomeric to RING4. RING11 is {gamma}-interferon inducible, a property shared with other genes involved in antigen presentation. Comparison between the amino acid sequences of RING11 and RING4 reveals strong homology. They propose that they form a heterodimer that transports peptides from the cytoplasm into the endoplasmic reticulum. They have identified two RING11 alleles, which differ in length of their derived protein sequence by 17 amino acids. The more common of these alleles is present in a Caucasoid population at a frequency of 79%.

  14. Tissue-Specific Transcript Profiling for ABC Transporters in the Sequestering Larvae of the Phytophagous Leaf Beetle Chrysomela populi

    Science.gov (United States)

    Gretscher, René R.; Groth, Marco; Boland, Wilhelm; Burse, Antje

    2014-01-01

    Background Insects evolved ingenious adaptations to use extraordinary food sources. Particularly, the diet of herbivores enriched with noxious plant secondary metabolites requires detoxification mechanisms. Sequestration, which involves the uptake, transfer, and concentration of occasionally modified phytochemicals into specialized tissues or hemolymph, is one of the most successful detoxification strategies found in most insect orders. Due to the ability of ATP-binding cassette (ABC) carriers to transport a wide range of molecules including phytochemicals and xenobiotics, it is highly likely that they play a role in this sequestration process. To shed light on the role of ABC proteins in sequestration, we describe an inventory of putative ABC transporters in various tissues in the sequestering juvenile poplar leaf beetle, Chrysomela populi. Results In the transcriptome of C. populi, we predicted 65 ABC transporters. To link the proteins with a possible function, we performed comparative phylogenetic analyses with ABC transporters of other insects and of humans. While tissue-specific profiling of each ABC transporter subfamily suggests that ABCB, C and G influence the plant metabolite absorption in the gut, ABCC with 14 members is the preferred subfamily responsible for the excretion of these metabolites via Malpighian tubules. Moreover, salicin, which is sequestered from poplar plants, is translocated into the defensive glands for further deterrent production. In these glands and among all identified ABC transporters, an exceptionally high transcript level was observed only for Cpabc35 (Cpmrp). RNAi revealed the deficiency of other ABC pumps to compensate the function of CpABC35, demonstrating its key role during sequestration. Conclusion We provide the first comprehensive phylogenetic study of the ABC family in a phytophagous beetle species. RNA-seq data from different larval tissues propose the importance of ABC pumps to achieve a homeostasis of plant

  15. Tissue-specific transcript profiling for ABC transporters in the sequestering larvae of the phytophagous leaf beetle Chrysomela populi.

    Directory of Open Access Journals (Sweden)

    Anja S Strauss

    Full Text Available Insects evolved ingenious adaptations to use extraordinary food sources. Particularly, the diet of herbivores enriched with noxious plant secondary metabolites requires detoxification mechanisms. Sequestration, which involves the uptake, transfer, and concentration of occasionally modified phytochemicals into specialized tissues or hemolymph, is one of the most successful detoxification strategies found in most insect orders. Due to the ability of ATP-binding cassette (ABC carriers to transport a wide range of molecules including phytochemicals and xenobiotics, it is highly likely that they play a role in this sequestration process. To shed light on the role of ABC proteins in sequestration, we describe an inventory of putative ABC transporters in various tissues in the sequestering juvenile poplar leaf beetle, Chrysomela populi.In the transcriptome of C. populi, we predicted 65 ABC transporters. To link the proteins with a possible function, we performed comparative phylogenetic analyses with ABC transporters of other insects and of humans. While tissue-specific profiling of each ABC transporter subfamily suggests that ABCB, C and G influence the plant metabolite absorption in the gut, ABCC with 14 members is the preferred subfamily responsible for the excretion of these metabolites via Malpighian tubules. Moreover, salicin, which is sequestered from poplar plants, is translocated into the defensive glands for further deterrent production. In these glands and among all identified ABC transporters, an exceptionally high transcript level was observed only for Cpabc35 (Cpmrp. RNAi revealed the deficiency of other ABC pumps to compensate the function of CpABC35, demonstrating its key role during sequestration.We provide the first comprehensive phylogenetic study of the ABC family in a phytophagous beetle species. RNA-seq data from different larval tissues propose the importance of ABC pumps to achieve a homeostasis of plant-derived compounds and

  16. Tissue-specific transcript profiling for ABC transporters in the sequestering larvae of the phytophagous leaf beetle Chrysomela populi.

    Science.gov (United States)

    Strauss, Anja S; Wang, Ding; Stock, Magdalena; Gretscher, René R; Groth, Marco; Boland, Wilhelm; Burse, Antje

    2014-01-01

    Insects evolved ingenious adaptations to use extraordinary food sources. Particularly, the diet of herbivores enriched with noxious plant secondary metabolites requires detoxification mechanisms. Sequestration, which involves the uptake, transfer, and concentration of occasionally modified phytochemicals into specialized tissues or hemolymph, is one of the most successful detoxification strategies found in most insect orders. Due to the ability of ATP-binding cassette (ABC) carriers to transport a wide range of molecules including phytochemicals and xenobiotics, it is highly likely that they play a role in this sequestration process. To shed light on the role of ABC proteins in sequestration, we describe an inventory of putative ABC transporters in various tissues in the sequestering juvenile poplar leaf beetle, Chrysomela populi. In the transcriptome of C. populi, we predicted 65 ABC transporters. To link the proteins with a possible function, we performed comparative phylogenetic analyses with ABC transporters of other insects and of humans. While tissue-specific profiling of each ABC transporter subfamily suggests that ABCB, C and G influence the plant metabolite absorption in the gut, ABCC with 14 members is the preferred subfamily responsible for the excretion of these metabolites via Malpighian tubules. Moreover, salicin, which is sequestered from poplar plants, is translocated into the defensive glands for further deterrent production. In these glands and among all identified ABC transporters, an exceptionally high transcript level was observed only for Cpabc35 (Cpmrp). RNAi revealed the deficiency of other ABC pumps to compensate the function of CpABC35, demonstrating its key role during sequestration. We provide the first comprehensive phylogenetic study of the ABC family in a phytophagous beetle species. RNA-seq data from different larval tissues propose the importance of ABC pumps to achieve a homeostasis of plant-derived compounds and offer a basis for

  17. Glycolysis inhibition inactivates ABC transporters to restore drug sensitivity in malignant cells.

    Directory of Open Access Journals (Sweden)

    Ayako Nakano

    Full Text Available Cancer cells eventually acquire drug resistance largely via the aberrant expression of ATP-binding cassette (ABC transporters, ATP-dependent efflux pumps. Because cancer cells produce ATP mostly through glycolysis, in the present study we explored the effects of inhibiting glycolysis on the ABC transporter function and drug sensitivity of malignant cells. Inhibition of glycolysis by 3-bromopyruvate (3BrPA suppressed ATP production in malignant cells, and restored the retention of daunorubicin or mitoxantrone in ABC transporter-expressing, RPMI8226 (ABCG2, KG-1 (ABCB1 and HepG2 cells (ABCB1 and ABCG2. Interestingly, although side population (SP cells isolated from RPMI8226 cells exhibited higher levels of glycolysis with an increased expression of genes involved in the glycolytic pathway, 3BrPA abolished Hoechst 33342 exclusion in SP cells. 3BrPA also disrupted clonogenic capacity in malignant cell lines including RPMI8226, KG-1, and HepG2. Furthermore, 3BrPA restored cytotoxic effects of daunorubicin and doxorubicin on KG-1 and RPMI8226 cells, and markedly suppressed subcutaneous tumor growth in combination with doxorubicin in RPMI8226-implanted mice. These results collectively suggest that the inhibition of glycolysis is able to overcome drug resistance in ABC transporter-expressing malignant cells through the inactivation of ABC transporters and impairment of SP cells with enhanced glycolysis as well as clonogenic cells.

  18. The Role of Eukaryotic and Prokaryotic ABC Transporter Family in Failure of Chemotherapy.

    Science.gov (United States)

    El-Awady, Raafat; Saleh, Ekram; Hashim, Amna; Soliman, Nehal; Dallah, Alaa; Elrasheed, Azza; Elakraa, Ghada

    2016-01-01

    Over the years chemotherapy failure has been a vital research topic as researchers have been striving to discover reasons behind it. The extensive studies carried out on chemotherapeutic agents confirm that resistance to chemotherapy is a major reason for treatment failure. "Resistance to chemotherapy," however, is a comprehensive phrase that refers to a variety of different mechanisms in which ATP-binding cassette (ABC) mediated efflux dominates. The ABC is one of the largest gene superfamily of transporters among both eukaryotes and prokaryotes; it represents a variety of genes that code for proteins, which perform countless functions, including drug efflux - a natural process that protects cells from foreign chemicals. Up to date, chemotherapy failure due to ABC drug efflux is an active research topic that continuously provides further evidence on multiple drug resistance (MDR), aiding scientists in tackling and overcoming this issue. This review focuses on drug resistance by ABC efflux transporters in human, viral, parasitic, fungal and bacterial cells and highlights the importance of the MDR permeability glycoprotein being the mutual ABC transporter among all studied organisms. Current developments and future directions to overcome this problem are also discussed.

  19. Complete inventory of ABC proteins in human pathogenic yeast, Candida albicans.

    Science.gov (United States)

    Gaur, Manisha; Choudhury, Devapriya; Prasad, Rajendra

    2005-01-01

    The recent completion of the sequencing project of the opportunistic human pathogenic yeast, Candida albicans (http://www.ncbi.nlm.nih.gov/), led us to analyze and classify its ATP-binding cassette (ABC) proteins, which constitute one of the largest superfamilies of proteins. Some of its members are multidrug transporters responsible for the commonly encountered problem of antifungal resistance. TBLASTN searches together with domain analysis identified 81 nucleotide-binding domains, which belong to 51 different putative open reading frames. Considering that each allelic pair represents a single ABC protein of the Candida genome, the total number of putative members of this superfamily is 28. Domain organization, sequence-based analysis and self-organizing map-based clustering led to the classification of Candida ABC proteins into 6 distinct subfamilies. Each subfamily from C. albicans has an equivalent in Saccharomyces cerevisiae suggesting a close evolutionary relationship between the two yeasts. Our searches also led to the identification of a new motif to each subfamily in Candida that could be used to identify sequences from the corresponding subfamily in other organisms. It is hoped that the inventory of Candida ABC transporters thus created will provide new insights into the role of ABC proteins in antifungal resistance as well as help in the functional characterization of the superfamily of these proteins. 2005 S. Karger AG, Basel

  20. Association of ATP-binding cassette transporter-A1 polymorphism ...

    Indian Academy of Sciences (India)

    ., Slagle S. and Eder H. A. 1982. Separation and quantitation of subclasses of human plasma high density lipoproteins by a simple precipitation procedure. J. Lipid. Res. 23, 1206–1223. Hallman D. M., Srinivasan S. R., Chen W., Boerwinkle E.

  1. Diversity of ABC transporter genes across the plant kingdom and their potential utility in biotechnology.

    Science.gov (United States)

    Lane, Thomas S; Rempe, Caroline S; Davitt, Jack; Staton, Margaret E; Peng, Yanhui; Soltis, Douglas Edward; Melkonian, Michael; Deyholos, Michael; Leebens-Mack, James H; Chase, Mark; Rothfels, Carl J; Stevenson, Dennis; Graham, Sean W; Yu, Jun; Liu, Tao; Pires, J Chris; Edger, Patrick P; Zhang, Yong; Xie, Yinlong; Zhu, Ying; Carpenter, Eric; Wong, Gane Ka-Shu; Stewart, C Neal

    2016-05-31

    The ATP-binding cassette (ABC) transporter gene superfamily is ubiquitous among extant organisms and prominently represented in plants. ABC transporters act to transport compounds across cellular membranes and are involved in a diverse range of biological processes. Thus, the applicability to biotechnology is vast, including cancer resistance in humans, drug resistance among vertebrates, and herbicide and other xenobiotic resistance in plants. In addition, plants appear to harbor the highest diversity of ABC transporter genes compared with any other group of organisms. This study applied transcriptome analysis to survey the kingdom-wide ABC transporter diversity in plants and suggest biotechnology applications of this diversity. We utilized sequence similarity-based informatics techniques to infer the identity of ABC transporter gene candidates from 1295 phylogenetically-diverse plant transcriptomes. A total of 97,149 putative (approximately 25 % were full-length) ABC transporter gene members were identified; each RNA-Seq library (plant sample) had 88 ± 30 gene members. As expected, simpler organisms, such as algae, had fewer unique members than vascular land plants. Differences were also noted in the richness of certain ABC transporter subfamilies. Land plants had more unique ABCB, ABCC, and ABCG transporter gene members on average (p plant groups (p plant groups. An increase in the number of gene family members present in the ABCB, ABCC, and ABCD transporter subfamilies may indicate an expansion of the ABC transporter superfamily among green land plants, which include all crop species. The striking difference between the number of ABCA subfamily transporter gene members between ferns and other plant taxa is surprising and merits further investigation. Discussed is the potential exploitation of ABC transporters in plant biotechnology, with an emphasis on crops.

  2. Expression pattern of the human ABC transporters in pluripotent embryonic stem cells and in their derivatives.

    Science.gov (United States)

    Erdei, Zsuzsa; Lőrincz, Réka; Szebényi, Kornélia; Péntek, Adrienn; Varga, Nóra; Likó, István; Várady, György; Szakács, Gergely; Orbán, Tamás I; Sarkadi, Balázs; Apáti, Agota

    2014-09-01

    ATP-binding cassette (ABC) transporters have key roles in various physiological functions as well as providing chemical defense and stress tolerance in human tissues. In this study, we have examined the expression pattern of all ABC proteins in pluripotent human embryonic stem cells (hESCs) and in their differentiated progenies. We paid special attention to the cellular expression and localization of multidrug transporter ABC proteins. Stem cell differentiation was carried out without chemical induction or cell sorting, and specialized cell types were separated mechanically. Cellular features regarding pluripotency and tissue identity, as well as ABC transporter expression were studied by flow cytomtery, immuno-microscopy, and qPCR-based low-density arrays. Pluripotent hESCs and differentiated cell types (cardiomyocytes, neuronal cells, and mesenchymal stem cells) were distinguished by morphology, immunostaining markers, and selected mRNA expression patterns. We found that the mRNA expression levels of the 48 human ABC proteins also clearly distinguished the pluripotent and the respective differentiated cell types. When multidrug and lipid transporter ABC protein expression was examined by using well characterized specific antibodies by flow cytometry and confocal microscopy, the protein expression data corresponded well to the mRNA expression results. Moreover, the cellular localization of these important human ABC transporter proteins could be established in the pluripotent and differentiated hESC derived samples. These studies provide valuable information regarding ABC protein expression in human stem cells and their differentiated offspring. The results may also help to obtain further information concerning the specialized cellular functions of selected ABC transporters. © 2014 Clinical Cytometry Society.

  3. The Schizosaccharomyces pombe mam1 gene encodes an ABC transporter mediating secretion of M-factor

    DEFF Research Database (Denmark)

    Christensen, P U; Davey, William John; Nielsen, O

    1997-01-01

    In the fission yeast Schizosaccharomyces pombe, cells of opposite mating type communicate via diffusible peptide pheromones prior to mating. We have cloned the S. pombe mam1 gene, which encodes a 1336-amino acid protein belonging to the ATP-binding cassette (ABC) superfamily. The mam1 gene is only...... expressed in M cells and the gene product is responsible for the secretion of the mating pheromone. M-factor, a nonapeptide that is S-farnesylated and carboxy-methylated on its C-terminal cysteine residue. The predicted Mam1 protein is highly homologous to mammalian multiple drug-resistance proteins...... and to the Saccharomyces cerevisiae STE6 gene product, which mediates export of a-factor mating pheromone. We show that STE6 can also mediate secretion of M-factor in S. pombe....

  4. Multi-Drug Resistance ABC Transporter Inhibition Enhances Murine Ventral Prostate Stem/Progenitor Cell Differentiation.

    Science.gov (United States)

    Samant, Mugdha D; Jackson, Courtney M; Felix, Carina L; Jones, Anthony J; Goodrich, David W; Foster, Barbara A; Huss, Wendy J

    2015-05-15

    Multi-drug resistance (MDR)-ATP binding cassette (ABC) transporters, ABCB1, ABCC1, and ABCG2 participate in the efflux of steroid hormones, estrogens, and androgens, which regulate prostate development and differentiation. The role of MDR-ABC efflux transporters in prostate epithelial proliferation and differentiation remains unclear. We hypothesized that MDR-ABC transporters regulate prostate differentiation and epithelium regeneration. Prostate epithelial differentiation was studied using histology, sphere formation assay, and prostate regeneration induced by cycles of repeated androgen withdrawal and replacement. Embryonic deletion of Abcg2 resulted in a decreased number of luminal cells in the prostate and increased sphere formation efficiency, indicating an imbalance in the prostate epithelial differentiation pattern. Decreased luminal cell number in the Abcg2 null prostate implies reduced differentiation. Enhanced sphere formation efficiency in Abcg2 null prostate cells implies activation of the stem/progenitor cells. Prostate regeneration was associated with profound activation of the stem/progenitor cells, indicating the role of Abcg2 in maintaining stem/progenitor cell pool. Since embryonic deletion of Abcg2 may result in compensation by other ABC transporters, pharmacological inhibition of MDR-ABC efflux was performed. Pharmacological inhibition of MDR-ABC efflux enhanced prostate epithelial differentiation in sphere culture and during prostate regeneration. In conclusion, Abcg2 deletion leads to activation of the stem/progenitor cells and enhances differentiating divisions; and pharmacological inhibition of MDR-ABC efflux leads to epithelial differentiation. Our study demonstrates for the first time that MDR-ABC efflux transporter inhibition results in enhanced prostate epithelial cell differentiation.

  5. Identification of ABC transporters acting in vitamin B12metabolism in Caenorhabditis elegans.

    Science.gov (United States)

    McDonald, Megan K; Fritz, Julie-Anne; Jia, Dongxin; Scheuchner, Deborah; Snyder, Floyd F; Stanislaus, Avalyn; Curle, Jared; Li, Liang; Stabler, Sally P; Allen, Robert H; Mains, Paul E; Gravel, Roy A

    2017-12-01

    Vitamin B 12 (cobalamin, Cbl) is a micronutrient essential to human health. Cbl is not utilized as is but must go through complex subcellular and metabolic processing to generate two cofactor forms: methyl-Cbl for methionine synthase, a cytosolic enzyme; and adenosyl-Cbl for methylmalonyl-CoA mutase, a mitochondrial enzyme. Some 10-12 human genes have been identified responsible for the intracellular conversion of Cbl to cofactor forms, including genes that code for ATP-binding cassette (ABC) transporters acting at the lysosomal and plasma membranes. However, the gene for mitochondrial uptake is not known. We hypothesized that ABC transporters should be candidates for other uptake and efflux functions, including mitochondrial transport, and set out to screen ABC transporter mutants for blocks in Cbl utilization using the nematode roundworm Caenorhabditis elegans. Thirty-seven mutant ABC transporters were screened for the excretion of methylmalonic acid (MMA), which should result from loss of Cbl transport into the mitochondria. One mutant, wht-6, showed elevated MMA excretion and reduced [ 14 C]-propionate incorporation, pointing to a functional block in methylmalonyl-CoA mutase. In contrast, the wht-6 mutant appeared to have a normal cytosolic pathway based on analysis of cystathionine excretion, suggesting that cytosolic methionine synthase was functioning properly. Further, the MMA excretion in wht-6 could be partially reversed by including vitamin B 12 in the assay medium. The human ortholog of wht-6 is a member of the G family of ABC transporters. We propose wht-6 as a candidate for the transport of Cbl into mitochondria and suggest that a member of the corresponding ABCG family of ABC transporters has this role in humans. Our ABC transporter screen also revealed that mrp-1 and mrp-2 mutants excreted lower MMA than wild type, suggesting they were concentrating intracellular Cbl, consistent with the cellular efflux defect proposed for the mammalian MRP1 ABC

  6. ABC transporters in the liver fluke Opisthorchis felineus.

    Science.gov (United States)

    Mordvinov, Viatcheslav A; Ershov, Nikita I; Pirozhkova, Darya S; Pakharukov, Yuri V; Pakharukova, Maria Y

    2017-09-01

    ATP-binding cassette transporters (ABC transporters) are essential components of normal cellular physiological machinery in all eukaryotic and prokaryotic species, including parasites. Some ABC transporters, e.g., P-glycoproteins, are involved in the efflux of toxins and xenobiotics from the cell. At present, nothing is known about ABC transporter genes in epidemiologically important liver flukes from the Opisthorchiidae family, including European liver fluke Opisthorchis felineus. Opisthorchiasis caused by O. felineus is a serious public health problem on the territory of Russia and other Eastern European countries. ABC drug transporters are attractive objects of research on molecular markers of resistance and on ways to potentiate sensitivity to anthelmintics through suppression of the transporters themselves with specific inhibitors. Here we aimed at the identification of ABC transporters in the O. felineus transcriptome and identification of P-glycoproteins. In addition, our aim was to assess ABC transcript abundance in the RNA-seq data, to study the mRNA expression of P-glycoprotein genes by Droplet Digital PCR throughout the life cycle of O. felineus, and to test the gene induction in response to xenobiotics or anthelminthic agents. We found 23 nucleotide sequences encoding ABC transporters belonging to different subfamilies, including four sequences of P-glycoproteins. According to the transcript abundance in the RNA-seq data, one of P-glycoproteins (P4) has the highest expression among all ABC genes in the adult worm. P-glycoproteins showed substantially differential mRNA expression throughout the fluke life cycle, with high expression in the adult worms. Putative activity of P-glycoproteins as xenobiotic efflux pumps was found to be linked to the excretory system of O. felineus and to be inhibited by verapamil or tariquidar. Thus, ABC drug transporters in the liver fluke O. felineus are functionally active, indicating that ABC drug transporters are likely

  7. ABC-F Proteins Mediate Antibiotic Resistance through Ribosomal Protection.

    Science.gov (United States)

    Sharkey, Liam K R; Edwards, Thomas A; O'Neill, Alex J

    2016-03-22

    Members of the ABC-F subfamily of ATP-binding cassette proteins mediate resistance to a broad array of clinically important antibiotic classes that target the ribosome of Gram-positive pathogens. The mechanism by which these proteins act has been a subject of long-standing controversy, with two competing hypotheses each having gained considerable support: antibiotic efflux versus ribosomal protection. Here, we report on studies employing a combination of bacteriological and biochemical techniques to unravel the mechanism of resistance of these proteins, and provide several lines of evidence that together offer clear support to the ribosomal protection hypothesis. Of particular note, we show that addition of purified ABC-F proteins to anin vitrotranslation assay prompts dose-dependent rescue of translation, and demonstrate that such proteins are capable of displacing antibiotic from the ribosomein vitro To our knowledge, these experiments constitute the first direct evidence that ABC-F proteins mediate antibiotic resistance through ribosomal protection.IMPORTANCEAntimicrobial resistance ranks among the greatest threats currently facing human health. Elucidation of the mechanisms by which microorganisms resist the effect of antibiotics is central to understanding the biology of this phenomenon and has the potential to inform the development of new drugs capable of blocking or circumventing resistance. Members of the ABC-F family, which includelsa(A),msr(A),optr(A), andvga(A), collectively yield resistance to a broader range of clinically significant antibiotic classes than any other family of resistance determinants, although their mechanism of action has been controversial since their discovery 25 years ago. Here we present the first direct evidence that proteins of the ABC-F family act to protect the bacterial ribosome from antibiotic-mediated inhibition. Copyright © 2016 Sharkey et al.

  8. Molecular characterization of ABC transporters in marine ciliate, Euplotes crassus: Identification and response to cadmium and benzo[a]pyrene.

    Science.gov (United States)

    Kim, Hokyun; Yim, Bora; Kim, Jisoo; Kim, Haeyeon; Lee, Young-Mi

    2017-11-30

    ATP-binding cassette (ABC) transporters participate in transporting various substances, including xenobiotics, in or out of cells. However, their genetic information and function in ciliates remain still unclear. In this study, we sequenced and characterized two ABC transporter genes (EcABCB and EcABCC), and investigated the effect of cadmium (Cd) and benzo[a]pyrene (B[a]P) on their function and gene expression, using efflux assay and real-time reverse transcription-polymerase chain reaction (qRT-PCR), respectively, in the marine ciliate, Euplotes crassus. Sequencing analysis and efflux assay showed that EcABCB and EcABCC are typical ABC transporters, possessing conserved function. Exposure to Cd (≥5mg/L) and B[a]P (≥50.5μg/L) enhanced accumulation of a substrate. A significant increase in the expression of EcABCB and EcABC mRNA was observed at lower concentration in response to Cd and B[a]P. Our findings indicate that Cd and B[a]P could inhibit the efflux function of ABC transporters, leading to cellular toxicity in the ciliate. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. The role of ABC transporters of the blood-brain barrier in opioid tolerance development

    Directory of Open Access Journals (Sweden)

    Kamila Środa-Pomianek

    2018-02-01

    Full Text Available Opioids constitute an important group of drugs used in chronic pain treatment, e.g. cancer pain. Unfortunately, the development of the organism’s tolerance to the analgesic activity of opioids, the tendency to develop addictions and undesirable side effects are the main causes reducing opioid efficiency in chronic pain treatment. Among many mechanisms connected to emerging of opioid resistance the ATP-binding cassette (ABC transporters present at the blood-brain barrier may play an important role. These transporter proteins, especially P-glycoprotein (ABCB1, MDR1, affect pharmacokinetics of many drugs and xenobiotics that are their substrates. ABC transporters reduce cellular uptake of drugs and/or increase their export from brain tissue to blood. Substrate specificity of P-glycoprotein is extremely wide and comprises many structurally and functionally unrelated compounds. What is interesting, substrate specificity of P-glycoprotein overlaps to a great extent the specificity of the isoforms of cytochrome P450 involved in drug metabolism. In the present review, the ABC proteins-mediated transport of opioids was discussed as well as the mechanisms of transport regulation. Cellular metabolism of various opioid drugs and the role of ABC transporters in their absorption, distribution and elimination were also described.

  10. Energy Coupling Efficiency in the Type I ABC Transporter GlnPQ.

    Science.gov (United States)

    Lycklama A Nijeholt, Jelger A; Vietrov, Ruslan; Schuurman-Wolters, Gea K; Poolman, Bert

    2018-03-16

    Solute transport via ATP binding cassette (ABC) importers involves receptor-mediated substrate binding, which is followed by ATP-driven translocation of the substrate across the membrane. How these steps are exactly initiated and coupled, and how much ATP it takes to complete a full transport cycle, are subject of debate. Here, we reconstitute the ABC importer GlnPQ in nanodiscs and in proteoliposomes and determine substrate-(in)dependent ATP hydrolysis and transmembrane transport. We determined the conformational states of the substrate-binding domains (SBDs) by single-molecule Förster resonance energy transfer measurements. We find that the basal ATPase activity (ATP hydrolysis in the absence of substrate) is mainly caused by the docking of the closed-unliganded state of the SBDs onto the transporter domain of GlnPQ and that, unlike glutamine, arginine binds both SBDs but does not trigger their closing. Furthermore, comparison of the ATPase activity in nanodiscs with glutamine transport in proteoliposomes shows that the stoichiometry of ATP per substrate is close to two. These findings help understand the mechanism of transport and the energy coupling efficiency in ABC transporters with covalently linked SBDs, which may aid our understanding of Type I ABC importers in general. Copyright © 2018 Elsevier Ltd. All rights reserved.

  11. A Critical View on ABC Transporters and Their Interacting Partners in Auxin Transport.

    Science.gov (United States)

    Geisler, Markus; Aryal, Bibek; di Donato, Martin; Hao, Pengchao

    2017-10-01

    Different subclasses of ATP-binding cassette (ABC) transporters have been implicated in the transport of native variants of the phytohormone auxin. Here, the putative, individual roles of key members belonging to the ABCB, ABCD and ABCG families, respectively, are highlighted and the knowledge of their assumed expression and transport routes is reviewed and compared with their mutant phenotypes. Protein-protein interactions between ABC transporters and regulatory components during auxin transport are summarized and their importance is critically discussed. There is a focus on the functional interaction between members of the ABCB family and the FKBP42, TWISTED DWARF1, acting as a chaperone during plasma membrane trafficking of ABCBs. Further, the mode and relevance of functional ABCB-PIN interactions is diagnostically re-evaluated. A new nomenclature describing precisely the most likely ABCB-PIN interaction scenarios is suggested. Finally, available tools for the detection and prediction of ABC transporter interactomes are summarized and the potential of future ABC transporter interactome maps is highlighted. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  12. The ABC transporter Rv1272c of Mycobacterium tuberculosis enhances the import of long-chain fatty acids in Escherichia coli.

    Science.gov (United States)

    Martin, Audrey; Daniel, Jaiyanth

    2018-02-05

    Mycobacterium tuberculosis (Mtb), which causes tuberculosis, is capable of accumulating triacylglycerol (TAG) by utilizing fatty acids from host cells. ATP-binding cassette (ABC) transporters are involved in transport processes in all organisms. Among the classical ABC transporters in Mtb none have been implicated in fatty acid import. Since the transport of fatty acids from the host cell is important for dormancy-associated TAG synthesis in the pathogen, mycobacterial ABC transporter(s) could potentially be involved in this process. Based on sequence identities with a bacterial ABC transporter that mediates fatty acid import for TAG synthesis, we identified Rv1272c, a hitherto uncharacterized ABC-transporter in Mtb that also shows sequence identities with a plant ABC transporter involved in fatty acid transport. We expressed Rv1272c in E. coli and show that it enhances the import of radiolabeled fatty acids. We also show that Rv1272c causes a significant increase in the metabolic incorporation of radiolabeled long-chain fatty acids into cardiolipin, a tetra-acylated phospholipid, and phosphatidylglycerol in E. coli. This is the first report on the function of Rv1272c showing that it displays a long-chain fatty acid transport function. Copyright © 2018 Elsevier Inc. All rights reserved.

  13. Chorioamnionitis Induces a Specific Signature of Placental ABC Transporters Associated with an Increase of miR-331-5p in the Human Preterm Placenta.

    Science.gov (United States)

    do Imperio, Guinever Eustaquio; Bloise, Enrrico; Javam, Mohsen; Lye, Phetcharawan; Constantinof, Andrea; Dunk, Caroline; Dos Reis, Fernando Marcos; Lye, Stephen James; Gibb, William; Ortiga-Carvalho, Tania M; Matthews, Stephen Giles

    2018-01-01

    The ATP-binding cassette (ABC) transporters mediate drug biodisposition and immunological responses in the placental barrier. In vitro infective challenges alter expression of specific placental ABC transporters. We hypothesized that chorioamnionitis induces a distinct pattern of ABC transporter expression. Gene expression of 50 ABC transporters was assessed using TaqMan® Human ABC Transporter Array, in preterm human placentas without (PTD; n=6) or with histological chorioamnionitis (PTDC; n=6). Validation was performed using qPCR, immunohistochemistry and Western blot. MicroRNAs known to regulate P-glycoprotein (P-gp) were examined by qPCR. Up-regulation of ABCB9, ABCC2 and ABCF2 mRNA was detected in chorioamnionitis (pABC transporters will likely lead to modified transport of clinically relevant compounds at the inflamed placenta. A better understanding of the potential role of these transporters in the events surrounding PTD may also enable new strategies to be developed for prevention and treatment of PTD. © 2018 The Author(s). Published by S. Karger AG, Basel.

  14. Beyond Competitive Inhibition: Regulation of ABC Transporters by Kinases and Protein-Protein Interactions as Potential Mechanisms of Drug-Drug Interactions.

    Science.gov (United States)

    Crawford, Rebecca R; Potukuchi, Praveen K; Schuetz, Erin G; Schuetz, John D

    2018-03-07

    ATP-binding cassette (ABC) transporters are transmembrane efflux transporters mediating the extrusion of an array of substrates ranging from amino acids and lipids to xenobiotics, and many therapeutic compounds, including anticancer drugs. The ABC transporters are also recognized as important contributors to pharmacokinetics, especially in drug-drug interactions and adverse drug effects. Drugs and xenobiotics, as well as pathological conditions, can influence the transcription of ABC transporters, or modify their activity or intracellular localization. Kinases can affect the aforementioned processes for ABC transporters as do protein interactions. In this review, we focus on the ABC transporters ABCB1, ABCB11, ABCC1, ABCC4 and ABCG2 and illustrate how kinases and protein-protein interactions affect these transporters. The clinical relevance of these factors is currently unknown, however these examples suggest that our understanding of drug-drug interactions will benefit from further knowledge of how kinases and protein-protein interactions affect ABC transporters. The American Society for Pharmacology and Experimental Therapeutics.

  15. Abc1: a new ABC transporter from the fission yeast Schizosaccharomyces pombe

    DEFF Research Database (Denmark)

    Christensen, P U; Davis, K; Nielsen, O

    1997-01-01

    We have isolated the abc1 gene from the fission yeast Schizosaccharomyces pombe. Sequence analysis suggests that the Abc1 protein is a member of the ABC superfamily of transporters and is composed of two structurally homologous halves, each consisting of a hydrophobic region of six transmembrane...... domains and a hydrophilic region containing one ATP-binding site. The abc1 gene appears to be expressed under all growth conditions but gene disruption experiments indicate that it is not essential for growth. The sequence of the abc1 gene has been deposited in the EMBL data library under the Accession...

  16. Frequent down-regulation of ABC transporter genes in prostate cancer

    International Nuclear Information System (INIS)

    Demidenko, Rita; Razanauskas, Deividas; Daniunaite, Kristina; Lazutka, Juozas Rimantas; Jankevicius, Feliksas; Jarmalaite, Sonata

    2015-01-01

    ATP-binding cassette (ABC) transporters are transmembrane proteins responsible for the efflux of a wide variety of substrates, including steroid metabolites, through the cellular membranes. For better characterization of the role of ABC transporters in prostate cancer (PCa) development, the profile of ABC transporter gene expression was analyzed in PCa and noncancerous prostate tissues (NPT). TaqMan Low Density Array (TLDA) human ABC transporter plates were used for the gene expression profiling in 10 PCa and 6 NPT specimens. ABCB1 transcript level was evaluated in a larger set of PCa cases (N = 78) and NPT (N = 15) by real-time PCR, the same PCa cases were assessed for the gene promoter hypermethylation by methylation-specific PCR. Expression of eight ABC transporter genes (ABCA8, ABCB1, ABCC6, ABCC9, ABCC10, ABCD2, ABCG2, and ABCG4) was significantly down-regulated in PCa as compared to NPT, and only two genes (ABCC4 and ABCG1) were up-regulated. Down-regulation of ABC transporter genes was prevalent in the TMPRSS2-ERG-negative cases. A detailed analysis of ABCB1 expression confirmed TLDA results: a reduced level of the transcript was identified in PCa in comparison to NPT (p = 0.048). Moreover, the TMPRSS2-ERG-negative PCa cases showed significantly lower expression of ABCB1 in comparison to NPT (p = 0.003) or the fusion-positive tumors (p = 0.002). Promoter methylation of ABCB1 predominantly occurred in PCa and was rarely detected in NPT (p < 0.001). The study suggests frequent down-regulation of the ABC transporter genes in PCa, especially in the TMPRSS2-ERG-negative tumors. The online version of this article (doi:10.1186/s12885-015-1689-8) contains supplementary material, which is available to authorized users

  17. Antagonistic changes in sensitivity to antifungal drugs by mutations of an important ABC transporter gene in a fungal pathogen.

    Directory of Open Access Journals (Sweden)

    Wenjun Guan

    2010-06-01

    Full Text Available Fungal pathogens can be lethal, especially among immunocompromised populations, such as patients with AIDS and recipients of tissue transplantation or chemotherapy. Prolonged usage of antifungal reagents can lead to drug resistance and treatment failure. Understanding mechanisms that underlie drug resistance by pathogenic microorganisms is thus vital for dealing with this emerging issue. In this study, we show that dramatic sequence changes in PDR5, an ABC (ATP-binding cassette efflux transporter protein gene in an opportunistic fungal pathogen, caused the organism to become hypersensitive to azole, a widely used antifungal drug. Surprisingly, the same mutations conferred growth advantages to the organism on polyenes, which are also commonly used antimycotics. Our results indicate that Pdr5p might be important for ergosterol homeostasis. The observed remarkable sequence divergence in the PDR5 gene in yeast strain YJM789 may represent an interesting case of adaptive loss of gene function with significant clinical implications.

  18. The substrate-binding protein in bacterial ABC transporters: dissecting roles in the evolution of substrate specificity.

    Science.gov (United States)

    Maqbool, Abbas; Horler, Richard S P; Muller, Axel; Wilkinson, Anthony J; Wilson, Keith S; Thomas, Gavin H

    2015-10-01

    ATP-binding cassette (ABC) transporters, although being ubiquitous in biology, often feature a subunit that is limited primarily to bacteria and archaea. This subunit, the substrate-binding protein (SBP), is a key determinant of the substrate specificity and high affinity of ABC uptake systems in these organisms. Most prokaryotes have many SBP-dependent ABC transporters that recognize a broad range of ligands from metal ions to amino acids, sugars and peptides. Herein, we review the structure and function of a number of more unusual SBPs, including an ABC transporter involved in the transport of rare furanose forms of sugars and an SBP that has evolved to specifically recognize the bacterial cell wall-derived murein tripeptide (Mtp). Both these examples illustrate that subtle changes in binding-site architecture, including changes in side chains not directly involved in ligand co-ordination, can result in significant alteration of substrate range in novel and unpredictable ways. © 2015 Authors; published by Portland Press Limited.

  19. Congruence ABC implies ABC

    OpenAIRE

    Ellenberg, Jordan S.

    2000-01-01

    The ABC conjecture of Masser and Oesterle' states that if (a,b,c) are coprime integers with a + b + c = 0, then sup(|a|,|b|,|c|) 0. Oesterle' has observed that if the ABC conjecture holds for all (a,b,c) with 16 | abc, then the full ABC conjecture holds. We extend that result to show that, for every integer N, the "congruence ABC conjecture" that ABC holds for all (a,b,c) with N|abc implies the full ABC conjecture.

  20. Surface charge-specific interactions between polymer nanoparticles and ABC transporters in Caco-2 cells

    Science.gov (United States)

    Bhattacharjee, Sourav; van Opstal, Edward J.; Alink, Gerrit M.; Marcelis, Antonius T. M.; Zuilhof, Han; Rietjens, Ivonne M. C. M.

    2013-06-01

    The surface charge-dependent transport of polymeric nanoparticles (PNPs) across Caco-2 monolayers grown on transwell culture systems as an in vitro model for intestinal transport was tested. The transport of well-characterized, monodisperse, and fluorescent tri-block copolymer nanoparticles (TCNPs/size 45 nm) and polystyrene nanoparticles (PSNPs/size 50 nm), with different surface charges (positive and negative), was quantified. The positive PNPs showed a higher intracellular uptake and flux across the Caco-2 monolayers than the negative PNPs. Multidrug resistance/P-glycoprotein (MDR1/P-gp), a specific ATP-binding cassette (ABC) transporter, was found to play a major role in the cellular efflux of positive PNPs, whereas the multidrug resistance protein 1 took part in the efflux of negative PNPs from Caco-2 cells. The positive PNPs also caused an increased cellular uptake and apical to basolateral transport of the carcinogen PhIP across the Caco-2 monolayer. The flavonoid quercetin, which is known to interact with ABC transporters, promoted the intracellular uptake of different PNPs and interfered with the normal distribution patterns of PNPs in the transwell system. These results indicate that PNPs display surface charge-specific interactions with ABC transporters and can even affect the bioavailability of toxic food-borne compounds (like pro-carcinogens).

  1. Surface charge-specific interactions between polymer nanoparticles and ABC transporters in Caco-2 cells

    Energy Technology Data Exchange (ETDEWEB)

    Bhattacharjee, Sourav, E-mail: sourav.bhattacharjee@wur.nl [Wageningen University, Laboratory of Organic Chemistry (Netherlands); Opstal, Edward J. van; Alink, Gerrit M. [Wageningen University, Division of Toxicology (Netherlands); Marcelis, Antonius T. M.; Zuilhof, Han [Wageningen University, Laboratory of Organic Chemistry (Netherlands); Rietjens, Ivonne M. C. M. [Wageningen University, Division of Toxicology (Netherlands)

    2013-06-15

    The surface charge-dependent transport of polymeric nanoparticles (PNPs) across Caco-2 monolayers grown on transwell culture systems as an in vitro model for intestinal transport was tested. The transport of well-characterized, monodisperse, and fluorescent tri-block copolymer nanoparticles (TCNPs/size {approx}45 nm) and polystyrene nanoparticles (PSNPs/size {approx}50 nm), with different surface charges (positive and negative), was quantified. The positive PNPs showed a higher intracellular uptake and flux across the Caco-2 monolayers than the negative PNPs. Multidrug resistance/P-glycoprotein (MDR1/P-gp), a specific ATP-binding cassette (ABC) transporter, was found to play a major role in the cellular efflux of positive PNPs, whereas the multidrug resistance protein 1 took part in the efflux of negative PNPs from Caco-2 cells. The positive PNPs also caused an increased cellular uptake and apical to basolateral transport of the carcinogen PhIP across the Caco-2 monolayer. The flavonoid quercetin, which is known to interact with ABC transporters, promoted the intracellular uptake of different PNPs and interfered with the normal distribution patterns of PNPs in the transwell system. These results indicate that PNPs display surface charge-specific interactions with ABC transporters and can even affect the bioavailability of toxic food-borne compounds (like pro-carcinogens).

  2. Surface charge-specific interactions between polymer nanoparticles and ABC transporters in Caco-2 cells

    International Nuclear Information System (INIS)

    Bhattacharjee, Sourav; Opstal, Edward J. van; Alink, Gerrit M.; Marcelis, Antonius T. M.; Zuilhof, Han; Rietjens, Ivonne M. C. M.

    2013-01-01

    The surface charge-dependent transport of polymeric nanoparticles (PNPs) across Caco-2 monolayers grown on transwell culture systems as an in vitro model for intestinal transport was tested. The transport of well-characterized, monodisperse, and fluorescent tri-block copolymer nanoparticles (TCNPs/size ∼45 nm) and polystyrene nanoparticles (PSNPs/size ∼50 nm), with different surface charges (positive and negative), was quantified. The positive PNPs showed a higher intracellular uptake and flux across the Caco-2 monolayers than the negative PNPs. Multidrug resistance/P-glycoprotein (MDR1/P-gp), a specific ATP-binding cassette (ABC) transporter, was found to play a major role in the cellular efflux of positive PNPs, whereas the multidrug resistance protein 1 took part in the efflux of negative PNPs from Caco-2 cells. The positive PNPs also caused an increased cellular uptake and apical to basolateral transport of the carcinogen PhIP across the Caco-2 monolayer. The flavonoid quercetin, which is known to interact with ABC transporters, promoted the intracellular uptake of different PNPs and interfered with the normal distribution patterns of PNPs in the transwell system. These results indicate that PNPs display surface charge-specific interactions with ABC transporters and can even affect the bioavailability of toxic food-borne compounds (like pro-carcinogens).

  3. Role of ABC transporters in veterinary medicine: pharmaco-toxicological implications.

    Science.gov (United States)

    Virkel, Guillermo; Ballent, Mariana; Lanusse, Carlos; Lifschitz, Adrian

    2018-01-31

    Unlike physicians, veterinary practitioners must deal with a number of animal species with crucial differences in anatomy, physiology and metabolism. Accordingly, the pharmacokinetic behaviour, the clinical efficacy and the adverse or toxic effects of drugs may differ across domestic animals. Moreover, the use of drugs in food-producing species may impose a risk for humans due to the generation of chemical residues in edible products, a major concern for public health and consumer's safety. As is clearly known in human beings, the ATP binding cassette (ABC) of transport proteins may influence the bioavailability and elimination of numerous drugs and other xenobiotics in domestic animals as well. A number of drugs, currently available in the veterinary market, are substrates of one or more transporters. Therefore, significant drug-drug interactions among ABC substrates may have unpredictable pharmaco-toxicological consequences in different species of veterinary interest. In this context, different investigations revealed the major relevance of P-gp and other transport proteins, like breast cancer resistance protein (BCRP) and multidrug resistance-associated proteins (MRPs), in both companion and livestock animals. Undoubtedly, the discovery of the ABC transporters and the deep understanding of their physiological role in the different species introduced a new paradigm into the veterinary pharmacology. This review focus on the expression and function of the major transport proteins expressed in species of veterinary interest, and their impact in drug disposition, efficacy and toxicity. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  4. Putative ABC transporter responsible for acetic acid resistance in Acetobacter aceti.

    Science.gov (United States)

    Nakano, Shigeru; Fukaya, Masahiro; Horinouchi, Sueharu

    2006-01-01

    Two-dimensional gel electrophoretic analysis of the membrane fraction of Acetobacter aceti revealed the presence of several proteins that were produced in response to acetic acid. A 60-kDa protein, named AatA, which was mostly induced by acetic acid, was prepared; aatA was cloned on the basis of its NH2-terminal amino acid sequence. AatA, consisting of 591 amino acids and containing ATP-binding cassette (ABC) sequences and ABC signature sequences, belonged to the ABC transporter superfamily. The aatA mutation with an insertion of the neomycin resistance gene within the aatA coding region showed reduced resistance to acetic acid, formic acid, propionic acid, and lactic acid, whereas the aatA mutation exerted no effects on resistance to various drugs, growth at low pH (adjusted with HCl), assimilation of acetic acid, or resistance to citric acid. Introduction of plasmid pABC101 containing aatA under the control of the Escherichia coli lac promoter into the aatA mutant restored the defect in acetic acid resistance. In addition, pABC101 conferred acetic acid resistance on E. coli. These findings showed that AatA was a putative ABC transporter conferring acetic acid resistance on the host cell. Southern blot analysis and subsequent nucleotide sequencing predicted the presence of aatA orthologues in a variety of acetic acid bacteria belonging to the genera Acetobacter and Gluconacetobacter. The fermentation with A. aceti containing aatA on a multicopy plasmid resulted in an increase in the final yield of acetic acid.

  5. The ABC of ECF transporters : discovery and initial characterization of ECF-type ATP-binding casette (ABC) importers

    NARCIS (Netherlands)

    ter Beek, Josy

    2012-01-01

    Josy ter Beek heeft een nieuwe klasse transporteiwitten in de celmembraan ontdekt en gekarakteriseerd. Aangezien deze transporter alleen door bacteriën wordt gebruikt en voor het transport van verscheidene belangrijke stoffen zorgt, kan informatie over deze nieuwe klasse transporters in de toekomst

  6. Salinomycin overcomes ABC transporter-mediated multidrug and apoptosis resistance in human leukemia stem cell-like KG-1a cells

    Energy Technology Data Exchange (ETDEWEB)

    Fuchs, Dominik [Research Group Molecular Neuro-Oncology, German Cancer Research Center, Im Neuenheimer Feld 280, D-69120 Heidelberg (Germany); Institute of Immunology, University of Heidelberg, Im Neuenheimer Feld 305, D-69120 Heidelberg (Germany); Daniel, Volker; Sadeghi, Mahmoud; Opelz, Gerhard [Institute of Immunology, University of Heidelberg, Im Neuenheimer Feld 305, D-69120 Heidelberg (Germany); Naujokat, Cord, E-mail: cord.naujokat@med.uni-heidelberg.de [Institute of Immunology, University of Heidelberg, Im Neuenheimer Feld 305, D-69120 Heidelberg (Germany)

    2010-04-16

    Leukemia stem cells are known to exhibit multidrug resistance by expression of ATP-binding cassette (ABC) transporters which constitute transmembrane proteins capable of exporting a wide variety of chemotherapeutic drugs from the cytosol. We show here that human promyeloblastic leukemia KG-1a cells exposed to the histone deacetylase inhibitor phenylbutyrate resemble many characteristics of leukemia stem cells, including expression of functional ABC transporters such as P-glycoprotein, BCRP and MRP8. Consequently, KG-1a cells display resistance to the induction of apoptosis by various chemotherapeutic drugs. Resistance to apoptosis induction by chemotherapeutic drugs can be reversed by cyclosporine A, which effectively inhibits the activity of P-glycoprotein and BCRP, thus demonstrating ABC transporter-mediated drug resistance in KG-1a cells. However, KG-1a are highly sensitive to apoptosis induction by salinomycin, a polyether ionophore antibiotic that has recently been shown to kill human breast cancer stem cell-like cells and to induce apoptosis in human cancer cells displaying multiple mechanisms of drug and apoptosis resistance. Whereas KG-1a cells can be adapted to proliferate in the presence of apoptosis-inducing concentrations of bortezomib and doxorubicin, salinomycin does not permit long-term adaptation of the cells to apoptosis-inducing concentrations. Thus, salinomycin should be regarded as a novel and effective agent for the elimination of leukemia stem cells and other tumor cells exhibiting ABC transporter-mediated multidrug resistance.

  7. In Vitro and Ex Vivo Model Systems to Measure ABC Transporter Activity at the Blood-Brain Barrier.

    Science.gov (United States)

    de Wit, Nienke M; Kooij, Gijs; de Vries, Helga E

    2016-01-01

    With the aging population the occurrence of central nervous system diseases such as cancer, mental disorders and neurodegenerative diseases, is expected to increase and hence, the demand for effective drugs. However, the passage of drugs across the blood-brain barrier represents a major challenge in accomplishing efficient brain delivery of therapeutic agents. This highly efficient barrier is composed of a monolayer of capillary endothelial cells supported by pericytes and astrocytic end-feet, that together effectively shield the brain from the blood. The brain microvascular endothelial cells form a physical and metabolic barrier where paracellular and transcellular transport of molecules in and out of the brain is closely regulated, allowing nutrients to pass but preventing the entry of harmful neurotoxic substances, including drugs. For this purpose brain endothelial cells express efficient efflux pumps, such as ATP binding cassette (ABC) transporters, which limit the delivery of drugs into the brain. To treat the above-mentioned chronic central nervous system disorders, it is crucial to design compounds that can pass the blood-brain barrier and thus the ABC transporters. In order to achieve this, representative models of the blood-brain barrier with predictive validity are necessary. This review discusses the current in vitro and ex vivo model systems that are used to measure ABC transporter activity in order to study potential in vivo efficacy of blood-brain barrier-drug passage.

  8. Rv2477c is an antibiotic-sensitive manganese-dependent ABC-F ATPase in Mycobacterium tuberculosis.

    Science.gov (United States)

    Daniel, Jaiyanth; Abraham, Liz; Martin, Amanda; Pablo, Xyryl; Reyes, Shelby

    2018-01-01

    The Rv2477c protein of Mycobacterium tuberculosis (Mtb) belongs to the ATP-binding cassette (ABC) subfamily F that contains proteins with tandem nucleotide-binding domains but lacking transmembrane domains. ABC-F subfamily proteins have been implicated in diverse cellular processes such as translation, antibiotic resistance, cell growth and nutrient sensing. In order to investigate the biochemical characteristics of Rv2477c, we expressed it in Escherichia coli, purified it and characterized its enzymatic functions. We show that Rv2477c displays strong ATPase activity (V max  = 45.5 nmol/mg/min; K m  = 90.5 μM) that is sensitive to orthovanadate. The ATPase activity was maximal in the presence of Mn 2+ at pH 5.2. The Rv2477c protein was also able to hydrolyze GTP, TTP and CTP but at lower rates. Glutamate to glutamine substitutions at amino acid residues 185 and 468 in the two Walker B motifs of Rv2477c severely inhibited its ATPase activity. The antibiotics tetracycline and erythromycin, which target protein translation, were able to inhibit the ATPase activity of Rv2477c. We postulate that Rv2477c could be involved in mycobacterial protein translation and in resistance to tetracyclines and macrolides. This is the first report of the biochemical characterization of an ABC-F subfamily protein in Mtb. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Exploring the ATP-binding site of P2X receptors

    Directory of Open Access Journals (Sweden)

    Thierry eChataigneau

    2013-12-01

    Full Text Available P2X receptors are ATP-gated non-selective cation channels involved in many different physiological processes, such as synaptic transmission, inflammation and neuropathic pain. They form homo- or heterotrimeric complexes and contain three ATP-binding sites in their extracellular domain. The recent determination of X-ray structures of a P2X receptor solved in two states, a resting closed state and an ATP-bound, open-channel state, has provided unprecedented information not only regarding the three-dimensional shape of the receptor, but also on putative conformational changes that couple ATP binding to channel opening. These data provide a structural template for interpreting the huge amount of functional, mutagenesis, and biochemical data collected during more than fifteen years. In particular, the interfacial location of the ATP binding site and ATP orientation have been successfully confirmed by these structural studies. It appears that ATP binds to inter-subunit cavities shaped like open jaws, whose tightening induces the opening of the ion channel. These structural data thus represent a firm basis for understanding the activation mechanism of P2X receptors.

  10. Molecular mechanism of ATP binding and ion channel activation in P2X receptors

    Energy Technology Data Exchange (ETDEWEB)

    Hattori, Motoyuki; Gouaux, Eric (Oregon HSU)

    2012-10-24

    P2X receptors are trimeric ATP-activated ion channels permeable to Na{sup +}, K{sup +} and Ca{sup 2+}. The seven P2X receptor subtypes are implicated in physiological processes that include modulation of synaptic transmission, contraction of smooth muscle, secretion of chemical transmitters and regulation of immune responses. Despite the importance of P2X receptors in cellular physiology, the three-dimensional composition of the ATP-binding site, the structural mechanism of ATP-dependent ion channel gating and the architecture of the open ion channel pore are unknown. Here we report the crystal structure of the zebrafish P2X4 receptor in complex with ATP and a new structure of the apo receptor. The agonist-bound structure reveals a previously unseen ATP-binding motif and an open ion channel pore. ATP binding induces cleft closure of the nucleotide-binding pocket, flexing of the lower body {beta}-sheet and a radial expansion of the extracellular vestibule. The structural widening of the extracellular vestibule is directly coupled to the opening of the ion channel pore by way of an iris-like expansion of the transmembrane helices. The structural delineation of the ATP-binding site and the ion channel pore, together with the conformational changes associated with ion channel gating, will stimulate development of new pharmacological agents.

  11. Nucleotide dependent packing differences in helical crystals of the ABC transporter MsbA.

    Science.gov (United States)

    Ward, Andrew; Mulligan, Sheila; Carragher, Bridget; Chang, Geoffrey; Milligan, Ronald A

    2009-03-01

    Bacterial ATP binding cassette (ABC) exporters fulfill a wide variety of transmembrane transport roles and are homologous to the human multidrug resistance P-glycoprotein. Recent X-ray structures of the exporters MsbA and Sav1866 have begun to describe the conformational changes that accompany the ABC transport cycle. Here we present cryo-electron microscopy structures of MsbA reconstituted into a lipid bilayer. Using ATPase inhibitors, we captured three nucleotide transition states of the transporter that were subsequently reconstituted into helical arrays. The enzyme-substrate complex (trapped by ADP-aluminum fluoride or AMPPNP) crystallized in a different helical lattice than the enzyme-product complex (trapped by ADP-vanadate). Approximately 20A resolution maps were calculated for each state and revealed MsbA to be a dimer with a large channel between the membrane spanning domains, similar to the outward facing crystal structures of MsbA and Sav1866. This suggests that while there are likely structural differences between the nucleotide transition states, membrane embedded MsbA remains in an outward facing conformation while nucleotide is bound.

  12. Structure of the MacAB-TolC ABC-type tripartite multidrug efflux pump

    Science.gov (United States)

    Llabrés, Salomé; Neuberger, Arthur; Blaza, James N.; Bai, Xiao-chen; Okada, Ui; Murakami, Satoshi; van Veen, Hendrik W.; Zachariae, Ulrich; Scheres, Sjors H.W.; Luisi, Ben F.

    2017-01-01

    The MacA-MacB-TolC assembly of Escherichia coli is a transmembrane machine that spans the cell envelope and actively extrudes substrates, including macrolide antibiotics and polypeptide virulence factors. These transport processes are energized by the ATPase MacB, a member of the ATP-binding cassette (ABC) superfamily. We present an electron cryo-microscopy structure of the ABC-type tripartite assembly at near-atomic resolution. A hexamer of the periplasmic protein MacA bridges between a TolC trimer in the outer membrane and a MacB dimer in the inner membrane, generating a quaternary structure with a central channel for substrate translocation. A gating ring found in MacA is proposed to act as a one-way valve in substrate transport. The MacB structure features an atypical transmembrane domain (TMD) with a closely packed dimer interface and a periplasmic opening that is the likely portal for substrate entry from the periplasm, with subsequent displacement through an allosteric transport mechanism. PMID:28504659

  13. The human ABC transporter pseudogene family: Evidence for transcription and gene-pseudogene interference.

    Science.gov (United States)

    Piehler, Armin P; Hellum, Marit; Wenzel, Jürgen J; Kaminski, Ellen; Haug, Kari Bente Foss; Kierulf, Peter; Kaminski, Wolfgang E

    2008-04-11

    Pseudogenes are an integral component of the human genome. Little attention, however, has so far been paid to the phenomenon that some pseudogenes are transcriptionally active. Recently, we demonstrated that the human ortholog of the rodent testis-specific ATP-binding cassette (ABC) transporter Abca17 is a ubiquitously transcribed pseudogene (ABCA17P). The aim of the present study was to establish a complete inventory of all ABC transporter pseudogenes in the human genome and to identify transcriptionally active ABC transporter pseudogenes. Moreover, we tested the hypothesis that a regulatory interdependency exists between ABC transporter pseudogenes and their parental protein coding equivalents. Systematic bioinformatic analysis revealed the existence of 22 ABC transporter pseudogenes within the human genome. We identified two clusters on chromosomes 15 and 16, respectively, which harbor almost half of all pseudogenes (n = 10). Available information from EST and mRNA databases and RT-PCR expression profiling indicate that a large portion of the ABC transporter pseudogenes (45%, n = 10) are transcriptionally active and some of them are expressed as alternative splice variants. We demonstrate that both pseudogenes of the pseudoxanthoma elasticum gene ABCC6, ABCC6P1 and ABCC6P2, are transcribed. ABCC6P1 and ABCC6 possess near-identical promoter sequences and their tissue-specific expression profiles are strikingly similar raising the possibility that they form a gene-pseudogene dual transcription unit. Intriguingly, targeted knockdown of the transcribed pseudogene ABCC6P1 resulted in a significant reduction of ABCC6 mRNA expression levels. The human genome contains a surprisingly small number of ABC transporter pseudogenes relative to other known gene families. They are unevenly distributed across the chromosomes. Importantly, a significant portion of the ABC transporter pseudogenes is transcriptionally active. The downregulation of ABCC6 mRNA levels by targeted

  14. The human ABC transporter pseudogene family: Evidence for transcription and gene-pseudogene interference

    Directory of Open Access Journals (Sweden)

    Haug Kari

    2008-04-01

    Full Text Available Abstract Background Pseudogenes are an integral component of the human genome. Little attention, however, has so far been paid to the phenomenon that some pseudogenes are transcriptionally active. Recently, we demonstrated that the human ortholog of the rodent testis-specific ATP-binding cassette (ABC transporter Abca17 is a ubiquitously transcribed pseudogene (ABCA17P. The aim of the present study was to establish a complete inventory of all ABC transporter pseudogenes in the human genome and to identify transcriptionally active ABC transporter pseudogenes. Moreover, we tested the hypothesis that a regulatory interdependency exists between ABC transporter pseudogenes and their parental protein coding equivalents. Results Systematic bioinformatic analysis revealed the existence of 22 ABC transporter pseudogenes within the human genome. We identified two clusters on chromosomes 15 and 16, respectively, which harbor almost half of all pseudogenes (n = 10. Available information from EST and mRNA databases and RT-PCR expression profiling indicate that a large portion of the ABC transporter pseudogenes (45%, n = 10 are transcriptionally active and some of them are expressed as alternative splice variants. We demonstrate that both pseudogenes of the pseudoxanthoma elasticum gene ABCC6, ABCC6P1 and ABCC6P2, are transcribed. ABCC6P1 and ABCC6 possess near-identical promoter sequences and their tissue-specific expression profiles are strikingly similar raising the possibility that they form a gene-pseudogene dual transcription unit. Intriguingly, targeted knockdown of the transcribed pseudogene ABCC6P1 resulted in a significant reduction of ABCC6 mRNA expression levels. Conclusion The human genome contains a surprisingly small number of ABC transporter pseudogenes relative to other known gene families. They are unevenly distributed across the chromosomes. Importantly, a significant portion of the ABC transporter pseudogenes is transcriptionally active

  15. Function and regulation of ATP-binding cassette transport proteins involved in hepatobiliary transport (vol 12, pg 13, 2000)

    NARCIS (Netherlands)

    Hooiveld, GJEJ; van Montfoort, JE; Meijer, DKF; Muller, M

    Hepatobiliary transport of endogenous and exogenous compounds is mediated by the coordinated action of multiple transport systems present at the sinusoidal (basolateral) and canalicular (apical) membrane domains of hepatocytes. During the last few years many of these transporters have been cloned

  16. A multidrug ABC transporter with a taste for salt.

    Directory of Open Access Journals (Sweden)

    Saroj Velamakanni

    2009-07-01

    Full Text Available LmrA is a multidrug ATP-binding cassette (ABC transporter from Lactococcus lactis with no known physiological substrate, which can transport a wide range of chemotherapeutic agents and toxins from the cell. The protein can functionally replace the human homologue ABCB1 (also termed multidrug resistance P-glycoprotein MDR1 in lung fibroblast cells. Even though LmrA mediates ATP-dependent transport, it can use the proton-motive force to transport substrates, such as ethidium bromide, across the membrane by a reversible, H(+-dependent, secondary-active transport reaction. The mechanism and physiological context of this reaction are not known.We examined ion transport by LmrA in electrophysiological experiments and in transport studies using radioactive ions and fluorescent ion-selective probes. Here we show that LmrA itself can transport NaCl by a similar secondary-active mechanism as observed for ethidium bromide, by mediating apparent H(+-Na(+-Cl(- symport. Remarkably, LmrA activity significantly enhances survival of high-salt adapted lactococcal cells during ionic downshift.The observations on H(+-Na(+-Cl(- co-transport substantiate earlier suggestions of H(+-coupled transport by LmrA, and indicate a novel link between the activity of LmrA and salt stress. Our findings demonstrate the relevance of investigations into the bioenergetics of substrate translocation by ABC transporters for our understanding of fundamental mechanisms in this superfamily. This study represents the first use of electrophysiological techniques to analyze substrate transport by a purified multidrug transporter.

  17. Identification of Residues in the Lipopolysaccharide ABC Transporter That Coordinate ATPase Activity with Extractor Function.

    Science.gov (United States)

    Simpson, Brent W; Owens, Tristan W; Orabella, Matthew J; Davis, Rebecca M; May, Janine M; Trauger, Sunia A; Kahne, Daniel; Ruiz, Natividad

    2016-10-18

    The surface of most Gram-negative bacteria is covered with lipopolysaccharide (LPS), creating a permeability barrier against toxic molecules, including many antimicrobials. To assemble LPS on their surface, Gram-negative bacteria must extract newly synthesized LPS from the inner membrane, transport it across the aqueous periplasm, and translocate it across the outer membrane. The LptA to -G proteins assemble into a transenvelope complex that transports LPS from the inner membrane to the cell surface. The Lpt system powers LPS transport from the inner membrane by using a poorly characterized ATP-binding cassette system composed of the ATPase LptB and the transmembrane domains LptFG. Here, we characterize a cluster of residues in the groove region of LptB that is important for controlling LPS transport. We also provide the first functional characterization of LptFG and identify their coupling helices that interact with the LptB groove. Substitutions at conserved residues in these coupling helices compromise both the assembly and function of the LptB 2 FG complex. Defects in LPS transport conferred by alterations in the LptFG coupling helices can be rescued by changing a residue in LptB that is adjacent to functionally important residues in the groove region. This suppression is achieved by increasing the ATPase activity of the LptB 2 FG complex. Taken together, these data identify a specific binding site in LptB for the coupling helices of LptFG that is responsible for coupling of ATP hydrolysis by LptB with LptFG function to achieve LPS extraction. Lipopolysaccharide (LPS) is synthesized at the cytoplasmic membrane of Gram-negative bacteria and transported across several compartments to the cell surface, where it forms a barrier that protects these organisms from antibiotics. The LptB 2 FG proteins form an ATP-binding cassette (ABC) transporter that uses energy from ATP hydrolysis in the cytoplasm to facilitate extraction of LPS from the outer face of the

  18. Ion specificity and ionic strength dependence of the osmoregulatory ABC transporter OpuA

    NARCIS (Netherlands)

    Mahmood, N. A. B. Nik; Biemans-Oldehinkel, Esther; Patzlaff, Jason S.; Schuurman-Wolters, Gea K.; Poolman, Bert

    2006-01-01

    The ATPase subunit of the osmoregulatory ATP- binding cassette transporterOpuAfrom Lactococcus lactis has a C- terminal extension, the tandem cystathionine beta- synthase ( CBS) domain, which constitutes the sensor that allows the transporter to sense and respond to osmotic stress ( Biemans-

  19. How to move an amphipathic molecule across a lipid bilayer: different mechanisms for different ABC transporters?

    Science.gov (United States)

    Theodoulou, Frederica L; Carrier, David J; Schaedler, Theresia A; Baldwin, Stephen A; Baker, Alison

    2016-06-15

    Import of β-oxidation substrates into peroxisomes is mediated by ATP binding cassette (ABC) transporters belonging to subfamily D. In order to enter the β-oxidation pathway, fatty acids are activated by conversion to fatty acyl-CoA esters, a reaction which is catalysed by acyl-CoA synthetases (ACSs). Here, we present evidence for an unusual transport mechanism, in which fatty acyl-CoA substrates are accepted by ABC subclass D protein (ABCD) transporters, cleaved by the transporters during transit across the lipid bilayer to release CoA, and ultimately re-esterified in the peroxisome lumen by ACSs which interact with the transporter. We propose that this solves the biophysical problem of moving an amphipathic molecule across the peroxisomal membrane, since the intrinsic thioesterase activity of the transporter permits separate membrane translocation pathways for the hydrophobic fatty acid moiety and the polar CoA moiety. The cleavage/re-esterification mechanism also has the potential to control entry of disparate substrates into the β-oxidation pathway when coupled with distinct peroxisomal ACSs. A different solution to the movement of amphipathic molecules across a lipid bilayer is deployed by the bacterial lipid-linked oligosaccharide (LLO) flippase, PglK, in which the hydrophilic head group and the hydrophobic polyprenyl tail of the substrate are proposed to have distinct translocation pathways but are not chemically separated during transport. We discuss a speculative alternating access model for ABCD proteins based on the mammalian ABC transporter associated with antigen processing (TAP) and compare it to the novel mechanism suggested by the recent PglK crystal structures and biochemical data. © 2016 The Author(s).

  20. Conserved mechanisms of microtubule-stimulated ADP release, ATP binding, and force generation in transport kinesins.

    Science.gov (United States)

    Atherton, Joseph; Farabella, Irene; Yu, I-Mei; Rosenfeld, Steven S; Houdusse, Anne; Topf, Maya; Moores, Carolyn A

    2014-09-10

    Kinesins are a superfamily of microtubule-based ATP-powered motors, important for multiple, essential cellular functions. How microtubule binding stimulates their ATPase and controls force generation is not understood. To address this fundamental question, we visualized microtubule-bound kinesin-1 and kinesin-3 motor domains at multiple steps in their ATPase cycles--including their nucleotide-free states--at ∼ 7 Å resolution using cryo-electron microscopy. In both motors, microtubule binding promotes ordered conformations of conserved loops that stimulate ADP release, enhance microtubule affinity and prime the catalytic site for ATP binding. ATP binding causes only small shifts of these nucleotide-coordinating loops but induces large conformational changes elsewhere that allow force generation and neck linker docking towards the microtubule plus end. Family-specific differences across the kinesin-microtubule interface account for the distinctive properties of each motor. Our data thus provide evidence for a conserved ATP-driven mechanism for kinesins and reveal the critical mechanistic contribution of the microtubule interface.

  1. Cowpox virus protein CPXV012 eludes CTLs by blocking ATP binding to TAP.

    Science.gov (United States)

    Luteijn, Rutger D; Hoelen, Hanneke; Kruse, Elisabeth; van Leeuwen, Wouter F; Grootens, Jennine; Horst, Daniëlle; Koorengevel, Martijn; Drijfhout, Jan W; Kremmer, Elisabeth; Früh, Klaus; Neefjes, Jacques J; Killian, Antoinette; Lebbink, Robert Jan; Ressing, Maaike E; Wiertz, Emmanuel J H J

    2014-08-15

    CD8(+) CTLs detect virus-infected cells through recognition of virus-derived peptides presented at the cell surface by MHC class I molecules. The cowpox virus protein CPXV012 deprives the endoplasmic reticulum (ER) lumen of peptides for loading onto newly synthesized MHC class I molecules by inhibiting the transporter associated with Ag processing (TAP). This evasion strategy allows the virus to avoid detection by the immune system. In this article, we show that CPXV012, a 9-kDa type II transmembrane protein, prevents peptide transport by inhibiting ATP binding to TAP. We identified a segment within the ER-luminal domain of CPXV012 that imposes the block in peptide transport by TAP. Biophysical studies show that this domain has a strong affinity for phospholipids that are also abundant in the ER membrane. We discuss these findings in an evolutionary context and show that a frameshift deletion in the CPXV012 gene in an ancestral cowpox virus created the current form of CPXV012 that is capable of inhibiting TAP. In conclusion, our findings indicate that the ER-luminal domain of CPXV012 inserts into the ER membrane, where it interacts with TAP. CPXV012 presumably induces a conformational arrest that precludes ATP binding to TAP and, thus, activity of TAP, thereby preventing the presentation of viral peptides to CTLs. Copyright © 2014 by The American Association of Immunologists, Inc.

  2. Zinc and ATP Binding of the Hexameric AAA-ATPase PilF from Thermus thermophilus

    Science.gov (United States)

    Salzer, Ralf; Herzberg, Martin; Nies, Dietrich H.; Joos, Friederike; Rathmann, Barbara; Thielmann, Yvonne; Averhoff, Beate

    2014-01-01

    The traffic AAA-ATPase PilF is essential for pilus biogenesis and natural transformation of Thermus thermophilus HB27. Recently, we showed that PilF forms hexameric complexes containing six zinc atoms coordinated by conserved tetracysteine motifs. Here we report that zinc binding is essential for complex stability. However, zinc binding is neither required for pilus biogenesis nor natural transformation. A number of the mutants did not exhibit any pili during growth at 64 °C but still were transformable. This leads to the conclusion that type 4 pili and the DNA translocator are distinct systems. At lower growth temperatures (55 °C) the zinc-depleted multiple cysteine mutants were hyperpiliated but defective in pilus-mediated twitching motility. This provides evidence that zinc binding is essential for the role of PilF in pilus dynamics. Moreover, we found that zinc binding is essential for complex stability but dispensable for ATPase activity. In contrast to many polymerization ATPases from mesophilic bacteria, ATP binding is not required for PilF complex formation; however, it significantly increases complex stability. These data suggest that zinc and ATP binding increase complex stability that is important for functionality of PilF under extreme environmental conditions. PMID:25202014

  3. The role of ABCG-type ABC transporters in phytohormone transport.

    Science.gov (United States)

    Borghi, Lorenzo; Kang, Joohyun; Ko, Donghwi; Lee, Youngsook; Martinoia, Enrico

    2015-10-01

    Plant hormones (phytohormones) integrate endogenous and exogenous signals thus synchronizing plant growth with environmental and developmental changes. Similar to animals, phytohormones have distinct source and target tissues, hence controlled transport and focused targeting are required for their functions. Many evidences accumulated in the last years about the regulation of long-distance and directional transport of phytohormones. ATP-binding cassette (ABC) transporters turned out to play major roles in routing phytohormones not only in the plant body but also towards the outer environment. The ABCG-type proteins ABCG25 and ABCG40 are high affinity abscisic acid (ABA) transporters. ABCG14 is highly co-expressed with cytokinin biosynthesis and is the major root-to-shoot cytokinin transporter. Pleiotropic drug resistance1 (PDR1) from Petunia hybrida transports strigolactones (SLs) from the root tip to the plant shoot but also outside to the rhizosphere, where SLs are the main attractants to mycorrhizal fungi. Last but not least, ABCG36 and ABCG37 possibly play a dual role in coumarine and IBA transport. © 2015 Authors.

  4. Identification of Conserved ABC Importers Necessary for Intracellular Survival of Legionella pneumophila in Multiple Hosts

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    Amrita Lama

    2017-11-01

    Full Text Available It is established that the human pathogen Legionella pneumophila becomes significantly augmented for infection of macrophages after intracellular growth in amoebae when compared to like-strains cultivated in laboratory media. Based on this observation, we reasoned that the most critical virulence determinants of L.p. are expressed by responding to stimuli generated by the protozoan host specifically; a process we term “protozoan-priming.” We sought to identify L.p. virulence factors that were required for replication in amoebae in order to highlight the genes necessary for production of the most infectious form of the bacterium. Using a transposon mutagenesis screen, we successfully identified 12 insertions that produced bacteria severely attenuated for growth in amoebae, while retaining a functional Dot/Icm type IVb secretion system. Seven of these insertion mutants were found dispensable for growth in macrophages, revealing attractive therapeutic targets that reside upstream of the pathogen-human interface. Two candidates identified, lpg0730 and lpg0122 were required for survival and replication in amoebae and macrophage host cells. Both genes are conserved among numerous important human pathogenic bacteria that can persist or replicate in amoebae. Each gene encodes a component of an ATP binding cassette (ABC transport complex of unknown function. We demonstrate the lpg0730 ortholog in Francisella tularensis subsp. novicida to be essential for colonization of both protozoan and mammalian host cells, highlighting conserved survival mechanisms employed by bacteria that utilize protozoa as an environmental reservoir for replication.

  5. ABC Transporter Subfamily D: Distinct Differences in Behavior between ABCD1–3 and ABCD4 in Subcellular Localization, Function, and Human Disease

    Science.gov (United States)

    2016-01-01

    ATP-binding cassette (ABC) transporters are one of the largest families of membrane-bound proteins and transport a wide variety of substrates across both extra- and intracellular membranes. They play a critical role in maintaining cellular homeostasis. To date, four ABC transporters belonging to subfamily D have been identified. ABCD1–3 and ABCD4 are localized to peroxisomes and lysosomes, respectively. ABCD1 and ABCD2 are involved in the transport of long and very long chain fatty acids (VLCFA) or their CoA-derivatives into peroxisomes with different substrate specificities, while ABCD3 is involved in the transport of branched chain acyl-CoA into peroxisomes. On the other hand, ABCD4 is deduced to take part in the transport of vitamin B12 from lysosomes into the cytosol. It is well known that the dysfunction of ABCD1 results in X-linked adrenoleukodystrophy, a severe neurodegenerative disease. Recently, it is reported that ABCD3 and ABCD4 are responsible for hepatosplenomegaly and vitamin B12 deficiency, respectively. In this review, the targeting mechanism and physiological functions of the ABCD transporters are summarized along with the related disease. PMID:27766264

  6. ABC Transporter Subfamily D: Distinct Differences in Behavior between ABCD1-3 and ABCD4 in Subcellular Localization, Function, and Human Disease.

    Science.gov (United States)

    Kawaguchi, Kosuke; Morita, Masashi

    2016-01-01

    ATP-binding cassette (ABC) transporters are one of the largest families of membrane-bound proteins and transport a wide variety of substrates across both extra- and intracellular membranes. They play a critical role in maintaining cellular homeostasis. To date, four ABC transporters belonging to subfamily D have been identified. ABCD1-3 and ABCD4 are localized to peroxisomes and lysosomes, respectively. ABCD1 and ABCD2 are involved in the transport of long and very long chain fatty acids (VLCFA) or their CoA-derivatives into peroxisomes with different substrate specificities, while ABCD3 is involved in the transport of branched chain acyl-CoA into peroxisomes. On the other hand, ABCD4 is deduced to take part in the transport of vitamin B 12 from lysosomes into the cytosol. It is well known that the dysfunction of ABCD1 results in X-linked adrenoleukodystrophy, a severe neurodegenerative disease. Recently, it is reported that ABCD3 and ABCD4 are responsible for hepatosplenomegaly and vitamin B 12 deficiency, respectively. In this review, the targeting mechanism and physiological functions of the ABCD transporters are summarized along with the related disease.

  7. ABC Transporter Subfamily D: Distinct Differences in Behavior between ABCD1–3 and ABCD4 in Subcellular Localization, Function, and Human Disease

    Directory of Open Access Journals (Sweden)

    Kosuke Kawaguchi

    2016-01-01

    Full Text Available ATP-binding cassette (ABC transporters are one of the largest families of membrane-bound proteins and transport a wide variety of substrates across both extra- and intracellular membranes. They play a critical role in maintaining cellular homeostasis. To date, four ABC transporters belonging to subfamily D have been identified. ABCD1–3 and ABCD4 are localized to peroxisomes and lysosomes, respectively. ABCD1 and ABCD2 are involved in the transport of long and very long chain fatty acids (VLCFA or their CoA-derivatives into peroxisomes with different substrate specificities, while ABCD3 is involved in the transport of branched chain acyl-CoA into peroxisomes. On the other hand, ABCD4 is deduced to take part in the transport of vitamin B12 from lysosomes into the cytosol. It is well known that the dysfunction of ABCD1 results in X-linked adrenoleukodystrophy, a severe neurodegenerative disease. Recently, it is reported that ABCD3 and ABCD4 are responsible for hepatosplenomegaly and vitamin B12 deficiency, respectively. In this review, the targeting mechanism and physiological functions of the ABCD transporters are summarized along with the related disease.

  8. Investigation of the quaternary structure of an ABC transporter in living cells using spectrally resolved resonance energy transfer

    Science.gov (United States)

    Singh, Deo Raj

    Forster resonance energy transfer (FRET) has become an important tool to study proteins inside living cells. It has been used to explore membrane protein folding and dynamics, determine stoichiometry and geometry of protein complexes, and measure the distance between two molecules. In this dissertation, we use a method based on FRET and optical micro-spectroscopy (OptiMiS) technology, developed in our lab, to probe the structure of dynamic (as opposed to static) protein complexes in living cells. We use this method to determine the association stoichiometry and quaternary structure of an ABC transporter in living cells. Specifically, the transporter we investigate originates from the pathogen Pseudomonas aeruginosa, which is a Gram-negative bacterium with several virulence factors, lipopolysaccharides being one of them. This pathogen coexpresses two unique forms of lipopolysaccharides on its surface, the A- and B-bands. The A-band polysaccharides, synthesized in the cytoplasm, are translocated into the periplasm through an ATP-binding-cassette (ABC) transporter consisting of a transmembranar protein, Wzm, and a nucleotide-binding protein, Wzt. In P. aeruginosa, all of the biochemical studies of A-band LPS are concentrated on the stages of the synthesis and ligation of polysaccharides (PSs), leaving the export stage involving ABC transporter unexplored. The mode of PS export through ABC transporters is still unknown. This difficulty is due to the lack of information about sub-unit composition and structure of this bi-component ABC transporter. Using the FRET-OptiMiS combination method developed by our lab, we found that Wzt forms a rhombus-shaped homo-tetramer which becomes a square upon co-expression with Wzm, and that Wzm forms a square-shaped homo-tetramer both in the presence and absence of Wzt. Based on these results, we propose a structural model for the double-tetramer complex formed by the bi-component ABC transporter in living cells. An understanding of the

  9. The peroxisomal ABC transporter family

    NARCIS (Netherlands)

    Wanders, Ronald J. A.; Visser, Wouter F.; van Roermund, Carlo W. T.; Kemp, Stephan; Waterham, Hans R.

    2007-01-01

    This review describes the current state of knowledge about the ABCD family of peroxisomal half adenosine-triphosphate-binding cassette (ABC) transporters. ABCDs are predicted to be present in a variety of eukaryotic organisms, although at present, only ABCDs in the yeast Saccharomyces cerevisiae,

  10. Inhibition of the Human ABC Efflux Transporters P-gp and ...

    Science.gov (United States)

    High body burdens of polybrominated diphenyl ethers (PBDEs) in infants and young children have led to increased concern over their potential impact on human development. PBDE exposure can alter the expression of genes involved in thyroid homeostasis, including those of ATP-binding cassette (ABC) transporters, which mediate cellular xenobiotic efflux. However, little information exists on how PBDEs interact with ABC transporters such as P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP). The purpose of this study was to evaluate the interactions of 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47) and its hydroxylated metabolite 6-OH-BDE-47 with P-gp and BCRP, using human MDR1- and BCRP-expressing membrane vesicles and stably transfected NIH-3T3-MDR1 and MDCK-BCRP cells. In P-gp membranes, BDE-47 did not affect P-gp activity; however, 6-OH-BDE-47 inhibited P-gp activity at low µM concentrations (IC50 = 11.7 µM). In BCRP membranes, BDE-47 inhibited BCRP activity; however, 6-OH-BDE-47 was a stronger inhibitor [IC50 = 45.9 µM (BDE-47) vs. IC50 = 9.4 µM (6-OH-BDE-47)]. Intracellular concentrations of known P-gp and BCRP substrates [(3H)-paclitaxel and (3H)-prazosin, respectively] were significantly higher (indicating less efflux) in NIH-3T3-MDR1 and MDCK-BCRP cells in the presence of 6-OH-BDE-47, but not BDE-47. Collectively, our results indicate that the BDE-47 metabolite 6-OH-BDE-47 is an inhibitor of both P-gp and BCRP efflux activity.

  11. Co-Induction of a Glutathione-S-transferase, a Glutathione Transporter and an ABC Transporter in Maize by Xenobiotics

    Science.gov (United States)

    Liu, Zhiqian; Song, Xiaoyu; Li, Xuefeng; Wang, Chengju

    2012-01-01

    Glutathione conjugation reactions are one of the principal mechanisms that plants utilize to detoxify xenobiotics. The induction by four herbicides (2,4-D, atrazine, metolachlor and primisulfuron) and a herbicide safener (dichlormid) on the expression of three genes, ZmGST27, ZmGT1 and ZmMRP1, encoding respectively a glutathione-S-transferase, a glutathione transporter and an ATP-binding cassette (ABC) transporter was studied in maize. The results demonstrate that the inducing effect on gene expression varies with both chemicals and genes. The expression of ZmGST27 and ZmMRP1 was up-regulated by all five compounds, whereas that of ZmGT1 was increased by atrazine, metolachlor, primisulfuron and dichlormid, but not by 2,4-D. For all chemicals, the inducing effect was first detected on ZmGST27. The finding that ZmGT1 is activated alongside ZmGST27 and ZmMRP1 suggests that glutathione transporters are an important component in the xenobiotic detoxification system of plants. PMID:22792398

  12. RH421 binds into the ATP-binding site on the Na+/K+-ATPase.

    Science.gov (United States)

    Huličiak, Miroslav; Bazgier, Václav; Berka, Karel; Kubala, Martin

    2017-10-01

    The Na + /K + -ATPase plays a key role in ion transport across the plasma membrane of all animal cells. The voltage-sensitive styrylpyrimidium dye RH421 has been used in several laboratories for monitoring of Na + /K + -ATPase kinetics. It is known, that RH421 can interact with the enzyme and it can influence its activity at micromolar concentrations, but structural details of this interaction are only poorly understood. Experiments with isolated large cytoplasmic loop (C45) of Na + /K + -ATPase revealed that RH421 can interact with this part of the protein with dissociation constant 1μM. The Trp-to-RH421 FRET performed on six single-tryptophan mutants revealed that RH421 binds directly into the ATP-binding site. This conclusion was further supported by results from molecular docking, site-directed mutagenesis and by competitive experiments using ATP. Experiments with C45/DPPC mixture revealed that RH421 can bind to both C45 and lipids, but only the former interaction was influenced by the presence of ATP. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Cry11Aa Interacts with the ATP-Binding Protein from Culex quinquefasciatus To Improve the Toxicity.

    Science.gov (United States)

    Zhang, Lingling; Zhao, Guohui; Hu, Xiaohua; Liu, Jiannan; Li, Mingwei; Batool, Khadija; Chen, Mingfeng; Wang, Junxiang; Xu, Jin; Huang, Tianpei; Pan, Xiaohong; Xu, Lei; Yu, Xiao-Qiang; Guan, Xiong

    2017-12-20

    Cry11Aa displays high toxicity to the larvae of several mosquito species, including Aedes, Culex, and Anopheles. To study its binding characterization against Culex quinquefasciatus, Cry11Aa was purified and western blot results showed that Cry11Aa could bind successfully to the brush border membrane vesicles. To identify Cry11Aa-binding proteins in C. quinquefasciatus, a biotin-based protein pull-down experiment was performed and seven Cry11Aa-binding proteins were isolated from the midgut of C. quinquefasciatus larvae. Analysis of liquid chromatography-tandem mass spectrometry showed that one of the Cry11Aa-binding proteins is the ATP-binding domain 1 family member B. To investigate its binding property and effect on the toxicity of Cry11Aa, western blot, far-western blot, enzyme-linked immunosorbent assay, and bioassays of Cry11Aa in the presence and absence of the recombinant ATP-binding protein were performed. Our results showed that the ATP-binding protein interacted with Cry11Aa and increased the toxicity of Cry11Aa against C. quinquefasciatus. Our study suggests that midgut proteins other than the toxin receptors may modulate the toxicity of Cry toxins against mosquitoes.

  14. Long-range coupling between ATP-binding and lever-arm regions in myosin via dielectric allostery

    Science.gov (United States)

    Sato, Takato; Ohnuki, Jun; Takano, Mitsunori

    2017-12-01

    A protein molecule is a dielectric substance, so the binding of a ligand is expected to induce dielectric response in the protein molecule, considering that ligands are charged or polar in general. We previously reported that binding of adenosine triphosphate (ATP) to molecular motor myosin actually induces such a dielectric response in myosin due to the net negative charge of ATP. By this dielectric response, referred to as "dielectric allostery," spatially separated two regions in myosin, the ATP-binding region and the actin-binding region, are allosterically coupled. In this study, from the statistically stringent analyses of the extensive molecular dynamics simulation data obtained in the ATP-free and the ATP-bound states, we show that there exists the dielectric allostery that transmits the signal of ATP binding toward the distant lever-arm region. The ATP-binding-induced electrostatic potential change observed on the surface of the main domain induced a movement of the converter subdomain from which the lever arm extends. The dielectric response was found to be caused by an underlying large-scale concerted rearrangement of the electrostatic bond network, in which highly conserved charged/polar residues are involved. Our study suggests the importance of the dielectric property for molecular machines in exerting their function.

  15. Thermodynamics of the ATPase cycle of GlcV, the nucleotide-binding domain of the glucose ABC transporter of Sulfolobus solfataricus

    NARCIS (Netherlands)

    Pretz, Monika G.; Albers, Sonja-Verena; Schuurman-Wolters, Gea; Tampe, Robert; Driessen, Arnold J. M.; van der Does, Chris

    2006-01-01

    ATP-binding cassette transporters drive the transport of substrates across the membrane by the hydrolysis of ATP. They typically have a conserved domain structure with two membrane-spanning domains that form the transport channel and two cytosolic nucleotide-binding domains ( NBDs) that energize the

  16. Live cell FRET microscopy: homo- and heterodimerization of two human peroxisomal ABC transporters, the adrenoleukodystrophy protein (ALDP, ABCD1) and PMP70 (ABCD3).

    Science.gov (United States)

    Hillebrand, Merle; Verrier, Sophie E; Ohlenbusch, Andreas; Schäfer, Annika; Söling, Hans-Dieter; Wouters, Fred S; Gärtner, Jutta

    2007-09-14

    The adrenoleukodystrophy protein (ALDP) and the 70-kDa peroxisomal membrane protein (PMP70) are half-ATP-binding cassette (ABC) transporters in the mammalian peroxisome membrane. Mutations in the gene encoding ALDP result in a devastating neurodegenerative disorder, X-linked adrenoleukodystrophy (X-ALD) that is associated with elevated levels of very long chain fatty acids because of impaired peroxisomal beta-oxidation. The interactions of peroxisomal ABC transporters, their role in the peroxisomal membrane, and their functions in disease pathogenesis are poorly understood. Studies on ABC transporters revealed that half-transporters have to dimerize to gain functionality. So far, conflicting observations are described for ALDP. By the use of in vitro methods (yeast two-hybrid and immunoprecipitation assays) on the one hand, it was shown that ALDP can form homodimers as well as heterodimers with PMP70 and ALDR, while on the other hand, it was demonstrated that ALDP and PMP70 exclusively homodimerize. To circumvent the problems of artificial interactions due to biochemical sample preparation in vitro, we investigated protein-protein interaction of ALDP in its physiological environment by FRET microscopy in intact living cells. The statistical relevance of FRET data was determined in two different ways using probability distribution shift analysis and Kolmogorov-Smirnov statistics. We demonstrate in vivo that ALDP and PMP70 form homodimers as well as ALDP/PMP70 heterodimers where ALDP homodimers predominate. Using C-terminal deletion constructs of ALDP, we demonstrate that the last 87 C-terminal amino acids harbor the most important protein domain mediating these interactions, and that the N-terminal transmembrane region of ALDP has an additional stabilization effect on ALDP homodimers. Loss of ALDP homo- or heterodimerization is highly relevant for understanding the disease mechanisms of X-ALD.

  17. Active transmembrane drug transport in microgravity: a validation study using an ABC transporter model [v1; ref status: indexed, http://f1000r.es/41n

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    Sergi Vaquer

    2014-08-01

    Full Text Available Abstract Microgravity has been shown to influence the expression of ABC (ATP-Binding Cassette transporters in bacteria, fungi and mammals, but also to modify the activity of certain cellular components with structural and functional similarities to ABC transporters. Changes in activity of ABC transporters could lead to important metabolic disorders and undesired pharmacological effects during spaceflights. However, no current means exist to study the functionality of these transporters in microgravity. To this end, a Vesicular Transport Assay® (Solvo Biotechnology, Hungary was adapted to evaluate multi-drug resistance-associated protein 2 (MRP2 trans-membrane estradiol-17-β-glucuronide (E17βG transport activity, when activated by adenosine-tri-phosphate (ATP during parabolic flights. Simple diffusion, ATP-independent transport and benzbromarone inhibition were also evaluated. A high accuracy engineering system was designed to perform, monitor and synchronize all procedures. Samples were analysed using a validated high sensitivity drug detection protocol. Experiments were performed in microgravity during parabolic flights, and compared to 1g on ground results using identical equipment and procedures in all cases. Our results revealed that sufficient equipment accuracy and analytical sensitivity were reached to detect transport activity in both gravitational conditions. Additionally, transport activity levels of on ground samples were within commercial transport standards, proving the validity of the methods and equipment used. MRP2 net transport activity was significantly reduced in microgravity, so was signal detected in simple diffusion samples. Ultra-structural changes induced by gravitational stress upon vesicle membranes or transporters could explain the current results, although alternative explanations are possible. Further research is needed to provide a conclusive answer in this regard. Nevertheless, the present validated technology

  18. Enterococcus faecalis Uses a Phosphotransferase System Permease and a Host Colonization-Related ABC Transporter for Maltodextrin Uptake.

    Science.gov (United States)

    Sauvageot, Nicolas; Mokhtari, Abdelhamid; Joyet, Philippe; Budin-Verneuil, Aurélie; Blancato, Víctor S; Repizo, Guillermo D; Henry, Céline; Pikis, Andreas; Thompson, John; Magni, Christian; Hartke, Axel; Deutscher, Josef

    2017-05-01

    Maltodextrin is a mixture of maltooligosaccharides, which are produced by the degradation of starch or glycogen. They are mostly composed of α-1,4- and some α-1,6-linked glucose residues. Genes presumed to code for the Enterococcus faecalis maltodextrin transporter were induced during enterococcal infection. We therefore carried out a detailed study of maltodextrin transport in this organism. Depending on their length (3 to 7 glucose residues), E. faecalis takes up maltodextrins either via MalT, a maltose-specific permease of the phosphoenolpyruvate (PEP):carbohydrate phosphotransferase system (PTS), or the ATP binding cassette (ABC) transporter MdxEFG-MsmX. Maltotriose, the smallest maltodextrin, is primarily transported by the PTS permease. A malT mutant therefore exhibits significantly reduced growth on maltose and maltotriose. The residual uptake of the trisaccharide is catalyzed by the ABC transporter, because a malT mdxF double mutant no longer grows on maltotriose. The trisaccharide arrives as maltotriose-6″-P in the cell. MapP, which dephosphorylates maltose-6'-P, also releases P i from maltotriose-6″-P. Maltotetraose and longer maltodextrins are mainly (or exclusively) taken up via the ABC transporter, because inactivation of the membrane protein MdxF prevents growth on maltotetraose and longer maltodextrins up to at least maltoheptaose. E. faecalis also utilizes panose and isopanose, and we show for the first time, to our knowledge, that in contrast to maltotriose, its two isomers are primarily transported via the ABC transporter. We confirm that maltodextrin utilization via MdxEFG-MsmX affects the colonization capacity of E. faecalis , because inactivation of mdxF significantly reduced enterococcal colonization and/or survival in kidneys and liver of mice after intraperitoneal infection. IMPORTANCE Infections by enterococci, which are major health care-associated pathogens, are difficult to treat due to their increasing resistance to clinically

  19. The Crystal Structure of the YknZ Extracellular Domain of ABC Transporter YknWXYZ from Bacillus amyloliquefaciens.

    Directory of Open Access Journals (Sweden)

    Yongbin Xu

    Full Text Available Bacillus possesses the peptide toxin Sporulation-Delaying Protein (SDP, which can kill cells within a biofilm to support continued growth, thereby delaying the onset of biofilm sporulation. The four-component transporter YknWXYZ acts as a major SDP efflux pump to protect cells against the endogenous SDP toxin, for which YknYZ is a non-canonical ATP-binding cassette (ABC-type transporter. YknYZ consists of the following two components: (1 an individual protein (YknY and (2 a respective permease (YknZ. To date, the crystal structure, molecular function, and mechanism of action of the integral membrane protein YknZ remain to be elucidated. In this study, to characterize the structural and biochemical roles of YknZ in the functional assembly of YknWXYZ, we predicted and overexpressed the YknZ extracellular domain. We determined the crystal structure of B. amyloliquefaciens YknZ at a resolution of 2.0 Å. The structure revealed that the YknZ extracellular region exhibits significant structural similarity with the MacB periplasmic domain, which is a non-canonical ABC-type transporter in the tripartite macrolide-specific efflux pump in Gram-negative bacteria. We also found that the YknZ extracellular domain can directly bind to an extracellular component of YknX. This structural and biochemical study provides insights into the assembly of YknWXYZ, which may be relevant to understanding cannibalistic peptide toxin resistance in Bacillus and controlling bacterial growth.

  20. Preliminary time-of-flight neutron diffraction studies of Escherichia coli ABC transport receptor phosphate-binding protein at the Protein Crystallography Station.

    Science.gov (United States)

    Sippel, K H; Bacik, J; Quiocho, F A; Fisher, S Z

    2014-06-01

    Inorganic phosphate is an essential molecule for all known life. Organisms have developed many mechanisms to ensure an adequate supply, even in low-phosphate conditions. In prokaryotes phosphate transport is instigated by the phosphate-binding protein (PBP), the initial receptor for the ATP-binding cassette (ABC) phosphate transporter. In the crystal structure of the PBP-phosphate complex, the phosphate is completely desolvated and sequestered in a deep cleft and is bound by 13 hydrogen bonds: 12 to protein NH and OH donor groups and one to a carboxylate acceptor group. The carboxylate plays a key recognition role by accepting a phosphate hydrogen. PBP phosphate affinity is relatively consistent across a broad pH range, indicating the capacity to bind monobasic (H2PO4-) and dibasic (HPO4(2-)) phosphate; however, the mechanism by which it might accommodate the second hydrogen of monobasic phosphate is unclear. To answer this question, neutron diffraction studies were initiated. Large single crystals with a volume of 8 mm3 were grown and subjected to hydrogen/deuterium exchange. A 2.5 Å resolution data set was collected on the Protein Crystallography Station at the Los Alamos Neutron Science Center. Initial refinement of the neutron data shows significant nuclear density, and refinement is ongoing. This is the first report of a neutron study from this superfamily.

  1. RNA-seq analysis of the effect of kanamycin and the ABC transporter AtWBC19 on Arabidopsis thaliana seedlings reveals changes in metal content.

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    Ayalew Mentewab

    Full Text Available Plants are exposed to antibiotics produced by soil microorganisms, but little is known about their responses at the transcriptional level. Likewise, few endogenous mechanisms of antibiotic resistance have been reported. The Arabidopsis thaliana ATP Binding Cassette (ABC transporter AtWBC19 (ABCG19 is known to confer kanamycin resistance, but the exact mechanism of resistance is not well understood. Here we examined the transcriptomes of control seedlings and wbc19 mutant seedlings using RNA-seq analysis. Exposure to kanamycin indicated changes in the organization of the photosynthetic apparatus, metabolic fluxes and metal uptake. Elemental analysis showed a 60% and 80% reduction of iron uptake in control and wbc19 mutant seedlings respectively, upon exposure to kanamycin. The drop in iron content was accompanied by the upregulation of the gene encoding for FERRIC REDUCTION OXIDASE 6 (FRO6 in mutant seedlings but not by the differential expression of other transport genes known to be induced by iron deficiency. In addition, wbc19 mutants displayed a distinct expression profile in the absence of kanamycin. Most notably the expression of several zinc ion binding proteins, including ZINC TRANSPORTER 1 PRECURSOR (ZIP1 was increased, suggesting abnormal zinc uptake. Elemental analysis confirmed a 50% decrease of zinc content in wbc19 mutants. Thus, the antibiotic resistance gene WBC19 appears to also have a role in zinc uptake.

  2. Genetic variation in the proximal promoter of ABC and SLC superfamilies: liver and kidney specific expression and promoter activity predict variation.

    Directory of Open Access Journals (Sweden)

    Stephanie E Hesselson

    2009-09-01

    Full Text Available Membrane transporters play crucial roles in the cellular uptake and efflux of an array of small molecules including nutrients, environmental toxins, and many clinically used drugs. We hypothesized that common genetic variation in the proximal promoter regions of transporter genes contribute to observed variation in drug response. A total of 579 polymorphisms were identified in the proximal promoters (-250 to +50 bp and flanking 5' sequence of 107 transporters in the ATP Binding Cassette (ABC and Solute Carrier (SLC superfamilies in 272 DNA samples from ethnically diverse populations. Many transporter promoters contained multiple common polymorphisms. Using a sliding window analysis, we observed that, on average, nucleotide diversity (pi was lowest at approximately 300 bp upstream of the transcription start site, suggesting that this region may harbor important functional elements. The proximal promoters of transporters that were highly expressed in the liver had greater nucleotide diversity than those that were highly expressed in the kidney consistent with greater negative selective pressure on the promoters of kidney transporters. Twenty-one promoters were evaluated for activity using reporter assays. Greater nucleotide diversity was observed in promoters with strong activity compared to promoters with weak activity, suggesting that weak promoters are under more negative selective pressure than promoters with high activity. Collectively, these results suggest that the proximal promoter region of membrane transporters is rich in variation and that variants in these regions may play a role in interindividual variation in drug disposition and response.

  3. ABC transporter PEN3/PDR8/ABCG36 interacts with calmodulin that, like PEN3, is required for Arabidopsis nonhost resistance.

    Science.gov (United States)

    Campe, Ruth; Langenbach, Caspar; Leissing, Franz; Popescu, George V; Popescu, Sorina C; Goellner, Katharina; Beckers, Gerold J M; Conrath, Uwe

    2016-01-01

    Nonhost resistance (NHR) is the most prevalent form of plant immunity. In Arabidopsis, NHR requires membrane-localized ATP-binding cassette (ABC) transporter PENETRATION (PEN) 3. Upon perception of pathogen-associated molecular patterns, PEN3 becomes phosphorylated, suggestive of PEN3 regulation by post-translational modification. Here, we investigated the PEN3 protein interaction network. We probed the Arabidopsis protein microarray AtPMA-5000 with the N-terminal cytoplasmic domain of PEN3. Several of the proteins identified to interact with PEN3 in vitro represent cellular Ca(2+) sensors, including calmodulin (CaM) 3, CaM7 and several CaM-like proteins, pointing to the importance of Ca(2+) sensing to PEN3-mediated NHR. We demonstrated co-localization of PEN3 and CaM7, and we confirmed PEN3-CaM interaction in vitro and in vivo by PEN3 pull-down with CaM Sepharose, CaM overlay assay and bimolecular fluorescence complementation. We also show that just like in pen3, NHR to the nonadapted fungal pathogens Phakopsora pachyrhizi and Blumeria graminis f.sp. hordei is compromised in the Arabidopsis cam7 and pen3 cam7 mutants. Our study discloses CaM7 as a PEN3-interacting protein crucial to Arabidopsis NHR and emphasizes the importance of Ca(2+) sensing to plant immunity. © 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.

  4. NCBI nr-aa BLAST: CBRC-DNOV-01-1957 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-DNOV-01-1957 ref|NP_031461.1| ATP-binding cassette, sub-family D, member 1 [Mus musculus] sp|P48410|ABC...D1_MOUSE ATP-binding cassette sub-family D member 1 (Adrenoleukodystrophy protein)

  5. ATP binding and hydrolysis-driven rate-determining events in the RFC-catalyzed PCNA clamp loading reaction.

    Science.gov (United States)

    Sakato, Miho; Zhou, Yayan; Hingorani, Manju M

    2012-02-17

    The multi-subunit replication factor C (RFC) complex loads circular proliferating cell nuclear antigen (PCNA) clamps onto DNA where they serve as mobile tethers for polymerases and coordinate the functions of many other DNA metabolic proteins. The clamp loading reaction is complex, involving multiple components (RFC, PCNA, DNA, and ATP) and events (minimally: PCNA opening/closing, DNA binding/release, and ATP binding/hydrolysis) that yield a topologically linked clamp·DNA product in less than a second. Here, we report pre-steady-state measurements of several steps in the reaction catalyzed by Saccharomyces cerevisiae RFC and present a comprehensive kinetic model based on global analysis of the data. Highlights of the reaction mechanism are that ATP binding to RFC initiates slow activation of the clamp loader, enabling it to open PCNA (at ~2 s(-1)) and bind primer-template DNA (ptDNA). Rapid binding of ptDNA leads to formation of the RFC·ATP·PCNA(open)·ptDNA complex, which catalyzes a burst of ATP hydrolysis. Another slow step in the reaction follows ATP hydrolysis and is associated with PCNA closure around ptDNA (8 s(-1)). Dissociation of PCNA·ptDNA from RFC leads to catalytic turnover. We propose that these early and late rate-determining events are intramolecular conformational changes in RFC and PCNA that control clamp opening and closure, and that ATP binding and hydrolysis switch RFC between conformations with high and low affinities, respectively, for open PCNA and ptDNA, and thus bookend the clamp loading reaction. Copyright © 2011 Elsevier Ltd. All rights reserved.

  6. Critical role of γ-phosphate in structural transition of Na,K-ATPase upon ATP binding

    Science.gov (United States)

    Petrushanko, Irina Yu.; Mitkevich, Vladimir A.; Anashkina, Anastasia A.; Klimanova, Elizaveta A.; Dergousova, Elena A.; Lopina, Olga D.; Makarov, Alexander A.

    2014-06-01

    Active transport of sodium and potassium ions by Na,K-ATPase is accompanied by the enzyme conformational transition between E1 and E2 states. ATP and ADP bind to Na,K-ATPase in the E1 conformation with similar affinity but the properties of enzyme in complexes with these nucleotides are different. We have studied thermodynamics of Na,K-ATPase binding with adenine nucleotides at different temperatures using isothermal titration calorimetry. Our data indicate that β-phosphate is involved in complex formation by increasing the affinity of adenine nucleotides to Na,K-ATPase by an order of magnitude, while γ-phosphate does not affect it. ATP binding to Na,K-ATPase in contrast to ADP binding generates a structural transition in the enzyme, which is consistent with the movement of a significant portion of the surface area to a solvent-protected state. We propose that ATP binding leads to convergence of the nucleotide-binding and phosphorylation domains transferring the enzyme from the ``E1-open'' to ``E1-closed'' conformation ready for phosphorylation.

  7. Professionsbachelorens ABC

    DEFF Research Database (Denmark)

    Svendsen, Christian Bundgaard

    2008-01-01

    På baggrund af Lars-Henrik Schmidts "Sociale ABC" og Bloom og Dreyfus' taksonomier gives der et teoretiske bud på professionsbachelorbegrebet......På baggrund af Lars-Henrik Schmidts "Sociale ABC" og Bloom og Dreyfus' taksonomier gives der et teoretiske bud på professionsbachelorbegrebet...

  8. Genetic association analysis of ATP binding cassette protein family reveals a novel association of ABCB1 genetic variants with epilepsy risk, but not with drug-resistance.

    Directory of Open Access Journals (Sweden)

    Shabeesh Balan

    Full Text Available Epilepsy constitutes a heterogeneous group of disorders that is characterized by recurrent unprovoked seizures due to widely different etiologies. Multidrug resistance remains a major issue in clinical epileptology, where one third of patients with epilepsy continue to have seizures. Role of efflux transporters in multidrug resistant epilepsy has been attributed to drug-resistant epilepsy although, with discrepant observation in genetic studies. These discrepancies could be attributed to variety of factors such as variable definition of the anti-epileptic drug (AED-resistance, variable epilepsy phenotypes and ethnicities among the studies. In the present study we inquired the role of multidrug transporters ABCB1 and ABCG2 variants in determining AED-resistance and susceptibility to epilepsy in three well-characterized cohorts comprising of mesial temporal lobe epilepsy with hippocampal sclerosis (MTLE-HS (prototype for AED-resistant epilepsy; juvenile myoclonic epilepsy (JME (prototype for AED-responsive epilepsy; and healthy non-epileptic controls, in 738 subjects of Malayalam speaking south Indian ancestry. ABCB1 and ABCG2 variants were not found to be associated with drug resistance when AED-resistant and AED-responsive cohorts were compared. However, a significant association was observed between ABCB1 (C3435T rs1045642 and risk of having epilepsy (MTLE-HS and JME pooled cohort; genotypic p-value = 0.0002; allelic p-value = 0.004. This association was seen persistent with MTLE-HS (genotypic p-value = 0.0008; allelic p-value = 0.004 and also with JME (genotypic p-value = 0.01; allelic p-value = 0.05 cohort individually. In-silico functional prediction indicated that ABCB1 rs1045642 has a deleterious impact on protein coding function and in splicing regulation. We conclude that the ABCB1 and ABCG2 variants do not confer to AED-resistance in the study population. However, ABCB1 rs1045642 increases vulnerability to epilepsy with greater tendency for MTLE-HS in south Indian ancestry from Kerala.

  9. Association of three common single nucleotide polymorphisms of ATP binding cassette G8 gene with gallstone disease: a meta-analysis.

    Directory of Open Access Journals (Sweden)

    Zhao-Yan Jiang

    Full Text Available In this study, we evaluated the association between these polymorphisms and gallstone disease using meta-analysis and compared the hepatic ABCG5/G8 mRNA expression and biliary lipids composition in patients with different genotypes of T400K and Y54C.Data were analyzed using the Stata/SE 11.0 software and a random- effects model was applied irrespective of between-study heterogeneity. Hepatic mRNA expression of ABCG5/G8 genes in 182 patients with gallstone disease and 35 gallstone-free patients who underwent cholecystectomy were determined using real-time PCR. Genotypes of Y54C and T400K in the ABCG8 gene were determined by allelic discrimination using either genomic DNA or hepatic cDNA as template by Taqman assays. Biliary compostion in gallbladder bile was assayed in these patients as well.Ten papers including 13 cohorts were included for the final analysis. In the genotype model, the overall association between genotype with gallstone was significant for D19H (OR = 2.43, 95%CI: 2.23-2.64, P<0.001, and for Y54C (OR = 1.36, 95%CI: 1.01-1.83, P = 0.044, or T400K (OR = 1.17, 95%CI: 0.96-1.43. P = 0.110. In allele model, minor alleles of D19H polymorphism (allele D: OR = 2.25, 95%CI: 2.10-2.42, P<0.001 and of T400K polymorphism (allele K: OR = 1.18, 95%CI: 1.06-1.31, P<0.001 were related with an increased risk of gallstone disease. However, minor allele of Y54C polymorphism (allele Y, OR = 1.08, 95%CI: 0.96-1.21, P = 0.146 was not related with gallstone disease. I(2 statistics indicated no significant between-study heterogeneity for all genetic models for any of the three polymorphisms. Funnel plot and Egger's test suggested the absence of publication bias as well. However, no association of T400K and Y54C polymorphism with hepatic ABCG8/G5 mRNA expression or biliary lipids composition was found.Our study showed strong association of D19H polymorphism with gallstone disease. T400K and Y54C polymorphism, though to a less extent, may also relate with gallstone disease.

  10. Scavenger Receptor Class B Type I Mediates Biliary Cholesterol Secretion Independent of ATP-Binding Cassette Transporter g5/g8 in Mice

    NARCIS (Netherlands)

    Wiersma, Harmen; Gatti, Alberto; Nijstad, Niels; Elferink, Ronald P. J. Oude; Kuipers, Folkert; Tietge, Uwe J. F.

    2009-01-01

    Scavenger receptor class B type I (SR-BI) mediates selective uptake of cholesterol from high-density lipoprotein (HDL) particles by the liver and influences biliary cholesterol secretion. However, it is not dear, if this effect is direct or indirect. The aim of this study was to determine the impact

  11. ATP-binding cassette transporters are enriched in non-caveolar detergent-insoluble glycosphingolipid-enriched membrane domains (DIGs) in human multidrug-resistant cancer cells

    NARCIS (Netherlands)

    Hinrichs, JWJ; Klappe, K; Hummel, [No Value; Kok, JW

    2004-01-01

    In this study we show that P-glycoprotein in multi-drug-resistant 2780AD human ovarian carcinoma cells and multidrug resistance-associated protein 1 in multi-drug-resistant HT29(col) human colon carcinoma cells are predominantly located in Lubrol-based detergent-insoluble glycosphingolipid-enriched

  12. Impact of genetic variants of ATP binding cassette B1, AICAR transformylase/IMP cyclohydrolase, folyl-polyglutamatesynthetase, and methylenetetrahydrofolatereductase on methotrexate toxicity.

    Science.gov (United States)

    Sala-Icardo, Luis; Lamana, Amalia; Ortiz, Ana María; García Lorenzo, Elena; Moreno Fresneda, Pablo; García-Vicuña, Rosario; González-Álvaro, Isidoro

    To analyze the effect of single nucleotide polymorphisms (SNPs) with well-known functional impact of methylenetetrahydrofolatereductase (MTHFR; rs1801131 and rs1801133), the membrane transporter ABCB1 (rs1045642), the AICAR transformylase/IMP cyclohydrolase (ATIC; rs2372536) and folyl-polyglutamatesynthetase (FPGS; rs1544105), on liver and bone marrow toxicity of methotrexate (MTX). We analyzed 1415 visits from 350 patients of the PEARL (Princesa Early Arthritis Register Longitudinal) study: (732 with MTX, 683 without MTX). The different SNPs were genotyped using specific TaqMan probes (Applied Biosystems). Multivariate analyzes were performed using generalized linear models in which the dependent variables were the levels of serum alanine aminotransferase (liver toxicity), leukocytes, platelets or hemoglobin (hematologic toxicity) and adjusted for clinical variables (disease activity, etc.), analytical (renal function, etc.), sociodemographic (age, sex, etc.) and genetic variants of MTHFR, ABCB1, ATIC and FPGS. The effect of these variables on the MTX doses prescribed throughout follow-up was also analyzed through multivariate analysis nested by visit and patient. When taking MTX, those patients carrying the CC genotype of rs1045642 in ABCB1 showed significantly higher GPT levels (7.1±2.0 U/L; P<.001). Carrying at least one G allele of rs1544105 in FPGS was associated with lower leukocyte (-0.67±0.32; 0.038), hemoglobin (-0.34±0.11g/dL; P=.002), and platelet (-11.8±4.7; P=.012) levels. The presence of the G allele of rs1544105 in FPGS, and the T allele of rs1801133 in MTHFR, was significantly associated with the use of lower doses of MTX. Our data suggest that genotyping functional variants in FGPS and MTHFR enzymes and the transporter ABCB1 could help to identify patients with increased risk of MTX toxicity. Copyright © 2016 Elsevier España, S.L.U. and Sociedad Española de Reumatología y Colegio Mexicano de Reumatología. All rights reserved.

  13. Tiaozhi Tongmai Granules reduce atherogenesis and promote the expression of ATP-binding cassette transporter A1 in rabbit atherosclerotic plaque macrophages and the liver

    Directory of Open Access Journals (Sweden)

    Qing Sun

    2014-07-01

    Conclusions: Tiaozhi Tongmai Granules appear to have an anti-atherogenic effect that is most likely mediated by simultaneously upregulating the protein expression of ABCA1 in rabbit atherosclerotic plaque macrophages and in the liver.

  14. Detergent screening and purification of the human liver ABC transporters BSEP (ABCB11 and MDR3 (ABCB4 expressed in the yeast Pichia pastoris.

    Directory of Open Access Journals (Sweden)

    Philipp Ellinger

    Full Text Available The human liver ATP-binding cassette (ABC transporters bile salt export pump (BSEP/ABCB11 and the multidrug resistance protein 3 (MDR3/ABCB4 fulfill the translocation of bile salts and phosphatidylcholine across the apical membrane of hepatocytes. In concert with ABCG5/G8, these two transporters are responsible for the formation of bile and mutations within these transporters can lead to severe hereditary diseases. In this study, we report the heterologous overexpression and purification of human BSEP and MDR3 as well as the expression of the corresponding C-terminal GFP-fusion proteins in the yeast Pichia pastoris. Confocal laser scanning microscopy revealed that BSEP-GFP and MDR3-GFP are localized in the plasma membrane of P. pastoris. Furthermore, we demonstrate the first purification of human BSEP and MDR3 yielding ∼1 mg and ∼6 mg per 100 g of wet cell weight, respectively. By screening over 100 detergents using a dot blot technique, we found that only zwitterionic, lipid-like detergents such as Fos-cholines or Cyclofos were able to extract both transporters in sufficient amounts for subsequent functional analysis. For MDR3, fluorescence-detection size exclusion chromatography (FSEC screens revealed that increasing the acyl chain length of Fos-Cholines improved monodispersity. BSEP purified in n-dodecyl-β-D-maltoside or Cymal-5 after solubilization with Fos-choline 16 from P. pastoris membranes showed binding to ATP-agarose. Furthermore, detergent-solubilized and purified MDR3 showed a substrate-inducible ATPase activity upon addition of phosphatidylcholine lipids. These results form the basis for further biochemical analysis of human BSEP and MDR3 to elucidate the function of these clinically relevant ABC transporters.

  15. Increased expression of the yeast multidrug resistance ABC transporter Pdr18 leads to increased ethanol tolerance and ethanol production in high gravity alcoholic fermentation

    Science.gov (United States)

    2012-01-01

    Background The understanding of the molecular basis of yeast tolerance to ethanol may guide the design of rational strategies to increase process performance in industrial alcoholic fermentations. A set of 21 genes encoding multidrug transporters from the ATP-Binding Cassette (ABC) Superfamily and Major Facilitator Superfamily (MFS) in S. cerevisiae were scrutinized for a role in ethanol stress resistance. Results A yeast multidrug resistance ABC transporter encoded by the PDR18 gene, proposed to play a role in the incorporation of ergosterol in the yeast plasma membrane, was found to confer resistance to growth inhibitory concentrations of ethanol. PDR18 expression was seen to contribute to decreased 3 H-ethanol intracellular concentrations and decreased plasma membrane permeabilization of yeast cells challenged with inhibitory ethanol concentrations. Given the increased tolerance to ethanol of cells expressing PDR18, the final concentration of ethanol produced during high gravity alcoholic fermentation by yeast cells devoid of PDR18 was lower than the final ethanol concentration produced by the corresponding parental strain. Moreover, an engineered yeast strain in which the PDR18 promoter was replaced in the genome by the stronger PDR5 promoter, leading to increased PDR18 mRNA levels during alcoholic fermentation, was able to attain a 6 % higher ethanol concentration and a 17 % higher ethanol production yield than the parental strain. The improved fermentative performance of yeast cells over-expressing PDR18 was found to correlate with their increased ethanol tolerance and ability to restrain plasma membrane permeabilization induced throughout high gravity fermentation. Conclusions PDR18 gene over-expression increases yeast ethanol tolerance and fermentation performance leading to the production of highly inhibitory concentrations of ethanol. PDR18 overexpression in industrial yeast strains appears to be a promising approach to improve alcoholic

  16. Detergent Screening and Purification of the Human Liver ABC Transporters BSEP (ABCB11) and MDR3 (ABCB4) Expressed in the Yeast Pichia pastoris

    Science.gov (United States)

    Stindt, Jan; Smits, Sander H. J.; Schmitt, Lutz

    2013-01-01

    The human liver ATP-binding cassette (ABC) transporters bile salt export pump (BSEP/ABCB11) and the multidrug resistance protein 3 (MDR3/ABCB4) fulfill the translocation of bile salts and phosphatidylcholine across the apical membrane of hepatocytes. In concert with ABCG5/G8, these two transporters are responsible for the formation of bile and mutations within these transporters can lead to severe hereditary diseases. In this study, we report the heterologous overexpression and purification of human BSEP and MDR3 as well as the expression of the corresponding C-terminal GFP-fusion proteins in the yeast Pichia pastoris. Confocal laser scanning microscopy revealed that BSEP-GFP and MDR3-GFP are localized in the plasma membrane of P. pastoris. Furthermore, we demonstrate the first purification of human BSEP and MDR3 yielding ∼1 mg and ∼6 mg per 100 g of wet cell weight, respectively. By screening over 100 detergents using a dot blot technique, we found that only zwitterionic, lipid-like detergents such as Fos-cholines or Cyclofos were able to extract both transporters in sufficient amounts for subsequent functional analysis. For MDR3, fluorescence-detection size exclusion chromatography (FSEC) screens revealed that increasing the acyl chain length of Fos-Cholines improved monodispersity. BSEP purified in n-dodecyl-β-D-maltoside or Cymal-5 after solubilization with Fos-choline 16 from P. pastoris membranes showed binding to ATP-agarose. Furthermore, detergent-solubilized and purified MDR3 showed a substrate-inducible ATPase activity upon addition of phosphatidylcholine lipids. These results form the basis for further biochemical analysis of human BSEP and MDR3 to elucidate the function of these clinically relevant ABC transporters. PMID:23593265

  17. Increased expression of the yeast multidrug resistance ABC transporter Pdr18 leads to increased ethanol tolerance and ethanol production in high gravity alcoholic fermentation.

    Science.gov (United States)

    Teixeira, Miguel C; Godinho, Cláudia P; Cabrito, Tânia R; Mira, Nuno P; Sá-Correia, Isabel

    2012-07-27

    The understanding of the molecular basis of yeast tolerance to ethanol may guide the design of rational strategies to increase process performance in industrial alcoholic fermentations. A set of 21 genes encoding multidrug transporters from the ATP-Binding Cassette (ABC) Superfamily and Major Facilitator Superfamily (MFS) in S. cerevisiae were scrutinized for a role in ethanol stress resistance. A yeast multidrug resistance ABC transporter encoded by the PDR18 gene, proposed to play a role in the incorporation of ergosterol in the yeast plasma membrane, was found to confer resistance to growth inhibitory concentrations of ethanol. PDR18 expression was seen to contribute to decreased ³H-ethanol intracellular concentrations and decreased plasma membrane permeabilization of yeast cells challenged with inhibitory ethanol concentrations. Given the increased tolerance to ethanol of cells expressing PDR18, the final concentration of ethanol produced during high gravity alcoholic fermentation by yeast cells devoid of PDR18 was lower than the final ethanol concentration produced by the corresponding parental strain. Moreover, an engineered yeast strain in which the PDR18 promoter was replaced in the genome by the stronger PDR5 promoter, leading to increased PDR18 mRNA levels during alcoholic fermentation, was able to attain a 6 % higher ethanol concentration and a 17 % higher ethanol production yield than the parental strain. The improved fermentative performance of yeast cells over-expressing PDR18 was found to correlate with their increased ethanol tolerance and ability to restrain plasma membrane permeabilization induced throughout high gravity fermentation. PDR18 gene over-expression increases yeast ethanol tolerance and fermentation performance leading to the production of highly inhibitory concentrations of ethanol. PDR18 overexpression in industrial yeast strains appears to be a promising approach to improve alcoholic fermentation performance for sustainable bio

  18. Increased expression of the yeast multidrug resistance ABC transporter Pdr18 leads to increased ethanol tolerance and ethanol production in high gravity alcoholic fermentation

    Directory of Open Access Journals (Sweden)

    Teixeira Miguel C

    2012-07-01

    Full Text Available Abstract Background The understanding of the molecular basis of yeast tolerance to ethanol may guide the design of rational strategies to increase process performance in industrial alcoholic fermentations. A set of 21 genes encoding multidrug transporters from the ATP-Binding Cassette (ABC Superfamily and Major Facilitator Superfamily (MFS in S. cerevisiae were scrutinized for a role in ethanol stress resistance. Results A yeast multidrug resistance ABC transporter encoded by the PDR18 gene, proposed to play a role in the incorporation of ergosterol in the yeast plasma membrane, was found to confer resistance to growth inhibitory concentrations of ethanol. PDR18 expression was seen to contribute to decreased 3 H-ethanol intracellular concentrations and decreased plasma membrane permeabilization of yeast cells challenged with inhibitory ethanol concentrations. Given the increased tolerance to ethanol of cells expressing PDR18, the final concentration of ethanol produced during high gravity alcoholic fermentation by yeast cells devoid of PDR18 was lower than the final ethanol concentration produced by the corresponding parental strain. Moreover, an engineered yeast strain in which the PDR18 promoter was replaced in the genome by the stronger PDR5 promoter, leading to increased PDR18 mRNA levels during alcoholic fermentation, was able to attain a 6 % higher ethanol concentration and a 17 % higher ethanol production yield than the parental strain. The improved fermentative performance of yeast cells over-expressing PDR18 was found to correlate with their increased ethanol tolerance and ability to restrain plasma membrane permeabilization induced throughout high gravity fermentation. Conclusions PDR18 gene over-expression increases yeast ethanol tolerance and fermentation performance leading to the production of highly inhibitory concentrations of ethanol. PDR18 overexpression in industrial yeast strains appears to be a promising approach to

  19. The Q Motif Is Involved in DNA Binding but Not ATP Binding in ChlR1 Helicase.

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    Hao Ding

    Full Text Available Helicases are molecular motors that couple the energy of ATP hydrolysis to the unwinding of structured DNA or RNA and chromatin remodeling. The conversion of energy derived from ATP hydrolysis into unwinding and remodeling is coordinated by seven sequence motifs (I, Ia, II, III, IV, V, and VI. The Q motif, consisting of nine amino acids (GFXXPXPIQ with an invariant glutamine (Q residue, has been identified in some, but not all helicases. Compared to the seven well-recognized conserved helicase motifs, the role of the Q motif is less acknowledged. Mutations in the human ChlR1 (DDX11 gene are associated with a unique genetic disorder known as Warsaw Breakage Syndrome, which is characterized by cellular defects in genome maintenance. To examine the roles of the Q motif in ChlR1 helicase, we performed site directed mutagenesis of glutamine to alanine at residue 23 in the Q motif of ChlR1. ChlR1 recombinant protein was overexpressed and purified from HEK293T cells. ChlR1-Q23A mutant abolished the helicase activity of ChlR1 and displayed reduced DNA binding ability. The mutant showed impaired ATPase activity but normal ATP binding. A thermal shift assay revealed that ChlR1-Q23A has a melting point value similar to ChlR1-WT. Partial proteolysis mapping demonstrated that ChlR1-WT and Q23A have a similar globular structure, although some subtle conformational differences in these two proteins are evident. Finally, we found ChlR1 exists and functions as a monomer in solution, which is different from FANCJ, in which the Q motif is involved in protein dimerization. Taken together, our results suggest that the Q motif is involved in DNA binding but not ATP binding in ChlR1 helicase.

  20. Mycobacterium tuberculosis Universal Stress Protein Rv2623 Regulates Bacillary Growth by ATP Binding: Requirement for Establishing Chronic Persistent Infection

    Energy Technology Data Exchange (ETDEWEB)

    Drumm, J.; Mi, K; Bilder, P; Sun, M; Lim, J; Bielefeldt-Ohmann, H; Basaraba, R; So, M; Zhu, G; et. al.

    2009-01-01

    Tuberculous latency and reactivation play a significant role in the pathogenesis of tuberculosis, yet the mechanisms that regulate these processes remain unclear. The Mycobacterium tuberculosisuniversal stress protein (USP) homolog, rv2623, is among the most highly induced genes when the tubercle bacillus is subjected to hypoxia and nitrosative stress, conditions thought to promote latency. Induction of rv2623 also occurs when M. tuberculosis encounters conditions associated with growth arrest, such as the intracellular milieu of macrophages and in the lungs of mice with chronic tuberculosis. Therefore, we tested the hypothesis that Rv2623 regulates tuberculosis latency. We observed that an Rv2623-deficient mutant fails to establish chronic tuberculous infection in guinea pigs and mice, exhibiting a hypervirulence phenotype associated with increased bacterial burden and mortality. Consistent with this in vivo growth-regulatory role, constitutive overexpression of rv2623 attenuates mycobacterial growth in vitro. Biochemical analysis of purified Rv2623 suggested that this mycobacterial USP binds ATP, and the 2.9-A-resolution crystal structure revealed that Rv2623 engages ATP in a novel nucleotide-binding pocket. Structure-guided mutagenesis yielded Rv2623 mutants with reduced ATP-binding capacity. Analysis of mycobacteria overexpressing these mutants revealed that the in vitro growth-inhibitory property of Rv2623 correlates with its ability to bind ATP. Together, the results indicate that i M. tuberculosis Rv2623 regulates mycobacterial growth in vitro and in vivo, and ii Rv2623 is required for the entry of the tubercle bacillus into the chronic phase of infection in the host; in addition, iii Rv2623 binds ATP; and iv the growth-regulatory attribute of this USP is dependent on its ATP-binding activity. We propose that Rv2623 may function as an ATP-dependent signaling intermediate in a pathway that promotes persistent infection.

  1. Direct ATP photolabeling of Escherichia coli recA proteins: identification of regions required for ATP binding

    International Nuclear Information System (INIS)

    Banks, G.R.; Sedgwick, S.G.

    1986-01-01

    When the Escherichia coli RecA protein is UV irradiated in the presence of [alpha- 32 P]ATP, a labeled protein--ATP adduct is formed. All the experimental evidence indicates that, in forming such an adduct, the ATP becomes specifically immobilized in the catalytically relevant ATP binding site. The adduct can also be identified after irradiation of E. coli cell lysates in a similar manner. This direct ATP photolabeling of RecA proteins has been used to identify regions of the polypeptide chain involved in the binding of ATP. The photolabeling of a RecA protein that lacks wild-type carboxy-terminal amino acids is not detectable. A RecA protein in which the amino-terminal sequence NH2-Ala-Ile-Asp-Glu-Asn- is replaced by NH2-Thr-Met-Ile-Thr-Asn-Ser-Ser-Ser- is only about 5% as efficiently photolabeled as the wild-type protein. Both of these RecA protein constructions, however, contain all the elements previously implicated, directly or indirectly, in the binding of ATP. ATP-photolabeled RecA protein has also been chemically cleaved at specific amino acids in order to identify regions of the polypeptide chain to which the nucleotide becomes covalently photolinked. The evidence is consistent with a region comprising amino acids 116-170. Thus, this work and that of others suggest that several disparate regions of the unfolded polypeptide chain may combine to form the ATP binding site upon protein folding or may influence binding through long-range effects

  2. Film sheet cassette

    International Nuclear Information System (INIS)

    1981-01-01

    A novel film sheet cassette is described for handling CAT photographic films under daylight conditions and facilitating their imaging. A detailed description of the design and operation of the cassette is given together with appropriate illustrations. The resulting cassette is a low-cost unit which is easily constructed and yet provides a sure light-tight seal for the interior contents of the cassette. The individual resilient fingers on the light-trap permit the ready removal of the slide plate for taking pictures. The stippled, non-electrostatic surface of the pressure plate ensures an air layer and free slidability of the film for removal and withdrawal of the film sheet. The advantage of the daylight system is that a darkroom need not be used for inserting and removing the film in and out of the cassette resulting in a considerable time saving. (U.K.)

  3. A lower isoelectric point increases signal sequence-mediated secretion of recombinant proteins through a bacterial ABC transporter.

    Science.gov (United States)

    Byun, Hyunjong; Park, Jiyeon; Kim, Sun Chang; Ahn, Jung Hoon

    2017-12-01

    Efficient protein production for industrial and academic purposes often involves engineering microorganisms to produce and secrete target proteins into the culture. Pseudomonas fluorescens has a TliDEF ATP-binding cassette transporter, a type I secretion system, which recognizes C-terminal LARD3 signal sequence of thermostable lipase TliA. Many proteins are secreted by TliDEF in vivo when recombined with LARD3, but there are still others that cannot be secreted by TliDEF even when LARD3 is attached. However, the factors that determine whether or not a recombinant protein can be secreted through TliDEF are still unknown. Here, we recombined LARD3 with several proteins and examined their secretion through TliDEF. We found that the proteins secreted via LARD3 are highly negatively charged with highly-acidic isoelectric points (pI) lower than 5.5. Attaching oligo-aspartate to lower the pI of negatively-charged recombinant proteins improved their secretion, and attaching oligo-arginine to negatively-charged proteins blocked their secretion by LARD3. In addition, negatively supercharged green fluorescent protein (GFP) showed improved secretion, whereas positively supercharged GFP did not secrete. These results disclosed that proteins' acidic pI and net negative charge are major factors that determine their secretion through TliDEF. Homology modeling for TliDEF revealed that TliD dimer forms evolutionarily-conserved positively-charged clusters in its pore and substrate entrance site, which also partially explains the pI dependence of the TliDEF-dependent secretions. In conclusion, lowering the isoelectric point improved LARD3-mediated protein secretion, both widening the range of protein targets for efficient production via secretion and signifying an important aspect of ABC transporter-mediated secretions. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  4. Identification of the magnesium-binding domain of the high affinity ATP binding-site of the Bacillus subtilis and Escherichia coli seca protein

    NARCIS (Netherlands)

    van der Wolk, J.P.W.; Klose, M; de Wit, Janny; Blaauwen, T.den; Freudl, R; Driessen, A.J.M.

    1995-01-01

    The homodimeric SecA protein is the peripheral subunit of the translocase, and couples the hydrolysis of ATP to the translocation of precursor proteins across the bacterial cytoplasmic membrane. The high affinity ATP binding activity of SecA resides in the amino-terminal domain of SecA. This domain

  5. Asymmetric switching in a homodimeric ABC transporter: a simulation study.

    Directory of Open Access Journals (Sweden)

    Jussi Aittoniemi

    2010-04-01

    Full Text Available ABC transporters are a large family of membrane proteins involved in a variety of cellular processes, including multidrug and tumor resistance and ion channel regulation. Advances in the structural and functional understanding of ABC transporters have revealed that hydrolysis at the two canonical nucleotide-binding sites (NBSs is co-operative and non-simultaneous. A conserved core architecture of bacterial and eukaryotic ABC exporters has been established, as exemplified by the crystal structure of the homodimeric multidrug exporter Sav1866. Currently, it is unclear how sequential ATP hydrolysis arises in a symmetric homodimeric transporter, since it implies at least transient asymmetry at the NBSs. We show by molecular dynamics simulation that the initially symmetric structure of Sav1866 readily undergoes asymmetric transitions at its NBSs in a pre-hydrolytic nucleotide configuration. MgATP-binding residues and a network of charged residues at the dimer interface are shown to form a sequence of putative molecular switches that allow ATP hydrolysis only at one NBS. We extend our findings to eukaryotic ABC exporters which often consist of two non-identical half-transporters, frequently with degeneracy substitutions at one of their two NBSs. Interestingly, many residues involved in asymmetric conformational switching in Sav1866 are substituted in degenerate eukaryotic NBS. This finding strengthens recent suggestions that the interplay of a consensus and a degenerate NBS in eukaroytic ABC proteins pre-determines the sequence of hydrolysis at the two NBSs.

  6. Selective RNA targeting and regulated signaling by RIG-I is controlled by coordination of RNA and ATP binding.

    Science.gov (United States)

    Fitzgerald, Megan E; Rawling, David C; Potapova, Olga; Ren, Xiaoming; Kohlway, Andrew; Pyle, Anna Marie

    2017-02-17

    RIG-I is an innate immune receptor that detects and responds to infection by deadly RNA viruses such as influenza, and Hepatitis C. In the cytoplasm, RIG-I is faced with a difficult challenge: it must sensitively detect viral RNA while ignoring the abundance of host RNA. It has been suggested that RIG-I has a ‘proof-reading’ mechanism for rejecting host RNA targets, and that disruptions of this selectivity filter give rise to autoimmune diseases. Here, we directly monitor RNA proof-reading by RIG-I and we show that it is controlled by a set of conserved amino acids that couple RNA and ATP binding to the protein (Motif III). Mutations of this motif directly modulate proof-reading by eliminating or enhancing selectivity for viral RNA, with major implications for autoimmune disease and cancer. More broadly, the results provide a physical explanation for the ATP-gated behavior of SF2 RNA helicases and receptor proteins.

  7. L1198F Mutation Resensitizes Crizotinib to ALK by Altering the Conformation of Inhibitor and ATP Binding Sites

    Directory of Open Access Journals (Sweden)

    Jian Li

    2017-02-01

    Full Text Available The efficacy of anaplastic lymphoma kinase (ALK positive non-small-cell lung cancer (NSCLC treatment with small molecule inhibitors is greatly challenged by acquired resistance. A recent study reported the newest generation inhibitor resistant mutation L1198F led to the resensitization to crizotinib, which is the first Food and Drug Administration (FDA approved drug for the treatment of ALK-positive NSCLC. It is of great importance to understand how this extremely rare event occurred for the purpose of overcoming the acquired resistance of such inhibitors. In this study, we exploited molecular dynamics (MD simulation to dissect the molecular mechanisms. Our MD results revealed that L1198F mutation of ALK resulted in the conformational change at the inhibitor site and altered the binding affinity of ALK to crizotinib and lorlatinib. L1198F mutation also affected the autoactivation of ALK as supported by the identification of His1124 and Tyr1278 as critical amino acids involved in ATP binding and phosphorylation. Our findings are valuable for designing more specific and potent inhibitors for the treatment of ALK-positive NSCLC and other types of cancer.

  8. L1198F Mutation Resensitizes Crizotinib to ALK by Altering the Conformation of Inhibitor and ATP Binding Sites.

    Science.gov (United States)

    Li, Jian; Sun, Rong; Wu, Yuehong; Song, Mingzhu; Li, Jia; Yang, Qianye; Chen, Xiaoyi; Bao, Jinku; Zhao, Qi

    2017-02-24

    The efficacy of anaplastic lymphoma kinase (ALK) positive non-small-cell lung cancer (NSCLC) treatment with small molecule inhibitors is greatly challenged by acquired resistance. A recent study reported the newest generation inhibitor resistant mutation L1198F led to the resensitization to crizotinib, which is the first Food and Drug Administration (FDA) approved drug for the treatment of ALK-positive NSCLC. It is of great importance to understand how this extremely rare event occurred for the purpose of overcoming the acquired resistance of such inhibitors. In this study, we exploited molecular dynamics (MD) simulation to dissect the molecular mechanisms. Our MD results revealed that L1198F mutation of ALK resulted in the conformational change at the inhibitor site and altered the binding affinity of ALK to crizotinib and lorlatinib. L1198F mutation also affected the autoactivation of ALK as supported by the identification of His1124 and Tyr1278 as critical amino acids involved in ATP binding and phosphorylation. Our findings are valuable for designing more specific and potent inhibitors for the treatment of ALK-positive NSCLC and other types of cancer.

  9. Mevalonate 5-diphosphate mediates ATP binding to the mevalonate diphosphate decarboxylase from the bacterial pathogen Enterococcus faecalis

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Chun-Liang; Mermoud, James C.; Paul, Lake N.; Steussy, Calvin Nicklaus; Stauffacher, Cynthia V. (Purdue)

    2017-10-12

    The mevalonate pathway produces isopentenyl diphosphate (IPP), a building block for polyisoprenoid synthesis, and is a crucial pathway for growth of the human bacterial pathogen Enterococcus faecalis. The final enzyme in this pathway, mevalonate diphosphate decarboxylase (MDD), acts on mevalonate diphosphate (MVAPP) to produce IPP while consuming ATP. This essential enzyme has been suggested as a therapeutic target for the treatment of drug-resistant bacterial infections. Here, we report functional and structural studies on the mevalonate diphosphate decarboxylase from E. faecalis (MDDEF). The MDDEF crystal structure in complex with ATP (MDDEF–ATP) revealed that the phosphate-binding loop (amino acids 97–105) is not involved in ATP binding and that the phosphate tail of ATP in this structure is in an outward-facing position pointing away from the active site. This suggested that binding of MDDEF to MVAPP is necessary to guide ATP into a catalytically favorable position. Enzymology experiments show that the MDDEF performs a sequential ordered bi-substrate reaction with MVAPP as the first substrate, consistent with the isothermal titration calorimetry (ITC) experiments. On the basis of ITC results, we propose that this initial prerequisite binding of MVAPP enhances ATP binding. In summary, our findings reveal a substrate-induced substrate-binding event that occurs during the MDDEF-catalyzed reaction. The disengagement of the phosphate-binding loop concomitant with the alternative ATP-binding configuration may provide the structural basis for antimicrobial design against these pathogenic enterococci.

  10. Insect Resistance to Bacillus thuringiensis Toxin Cry2Ab Is Conferred by Mutations in an ABC Transporter Subfamily A Protein.

    Directory of Open Access Journals (Sweden)

    Wee Tek Tay

    2015-11-01

    Full Text Available The use of conventional chemical insecticides and bacterial toxins to control lepidopteran pests of global agriculture has imposed significant selection pressure leading to the rapid evolution of insecticide resistance. Transgenic crops (e.g., cotton expressing the Bt Cry toxins are now used world wide to control these pests, including the highly polyphagous and invasive cotton bollworm Helicoverpa armigera. Since 2004, the Cry2Ab toxin has become widely used for controlling H. armigera, often used in combination with Cry1Ac to delay resistance evolution. Isolation of H. armigera and H. punctigera individuals heterozygous for Cry2Ab resistance in 2002 and 2004, respectively, allowed aspects of Cry2Ab resistance (level, fitness costs, genetic dominance, complementation tests to be characterised in both species. However, the gene identity and genetic changes conferring this resistance were unknown, as was the detailed Cry2Ab mode of action. No cross-resistance to Cry1Ac was observed in mutant lines. Biphasic linkage analysis of a Cry2Ab-resistant H. armigera family followed by exon-primed intron-crossing (EPIC marker mapping and candidate gene sequencing identified three independent resistance-associated INDEL mutations in an ATP-Binding Cassette (ABC transporter gene we named HaABCA2. A deletion mutation was also identified in the H. punctigera homolog from the resistant line. All mutations truncate the ABCA2 protein. Isolation of further Cry2Ab resistance alleles in the same gene from field H. armigera populations indicates unequal resistance allele frequencies and the potential for Bt resistance evolution. Identification of the gene involved in resistance as an ABC transporter of the A subfamily adds to the body of evidence on the crucial role this gene family plays in the mode of action of the Bt Cry toxins. The structural differences between the ABCA2, and that of the C subfamily required for Cry1Ac toxicity, indicate differences in the

  11. Detoxification of multiple heavy metals by a half-molecule ABC transporter, HMT-1, and coelomocytes of Caenorhabditis elegans.

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    Marc S Schwartz

    2010-03-01

    Full Text Available Developing methods for protecting organisms in metal-polluted environments is contingent upon our understanding of cellular detoxification mechanisms. In this regard, half-molecule ATP-binding cassette (ABC transporters of the HMT-1 subfamily are required for cadmium (Cd detoxification. HMTs have conserved structural architecture that distinguishes them from other ABC transporters and allows the identification of homologs in genomes of different species including humans. We recently discovered that HMT-1 from the simple, unicellular organism, Schizosaccharomyces pombe, SpHMT1, acts independently of phytochelatin synthase (PCS and detoxifies Cd, but not other heavy metals. Whether HMTs from multicellular organisms confer tolerance only to Cd or also to other heavy metals is not known.Using molecular genetics approaches and functional in vivo assays we showed that HMT-1 from a multicellular organism, Caenorhabditis elegans, functions distinctly from its S. pombe counterpart in that in addition to Cd it confers tolerance to arsenic (As and copper (Cu while acting independently of pcs-1. Further investigation of hmt-1 and pcs-1 revealed that these genes are expressed in different cell types, supporting the notion that hmt-1 and pcs-1 operate in distinct detoxification pathways. Interestingly, pcs-1 and hmt-1 are co-expressed in highly endocytic C. elegans cells with unknown function, the coelomocytes. By analyzing heavy metal and oxidative stress sensitivities of the coelomocyte-deficient C. elegans strain we discovered that coelomocytes are essential mainly for detoxification of heavy metals, but not of oxidative stress, a by-product of heavy metal toxicity.We established that HMT-1 from the multicellular organism confers tolerance to multiple heavy metals and is expressed in liver-like cells, the coelomocytes, as well as head neurons and intestinal cells, which are cell types that are affected by heavy metal poisoning in humans. We also showed

  12. Civilsamfundets ABC

    DEFF Research Database (Denmark)

    Lund, Anker Brink; Meyer, Gitte

    Filantropiske fonde og foreninger får stadig større indflydelse på udviklingen af det danske samfund – fra kunst, kultur, byggeri og byudvikling til forskning og udvikling af den danske velfærdsstat. Gennem 32 essays giver Anker Brink Lund og Gitte Meyer en oplysende – og polemisk – analyse af de...... for almenvelllet? CIVILSAMFUNDETS ABC er skrevet på baggrund af et udredningsprojekt om civilsamfundets rolle i det danske samfund. Såvel udredningsprojektet som ABC’en er finansieret af Realdania...

  13. Evolution of mal ABC transporter operons in the Thermococcales and Thermotogales

    Directory of Open Access Journals (Sweden)

    Gogarten J Peter

    2008-01-01

    Full Text Available Abstract Background The mal genes that encode maltose transporters have undergone extensive lateral transfer among ancestors of the archaea Thermococcus litoralis and Pyrococcus furiosus. Bacterial hyperthermophiles of the order Thermotogales live among these archaea and so may have shared in these transfers. The genome sequence of Thermotoga maritima bears evidence of extensive acquisition of archaeal genes, so its ancestors clearly had the capacity to do so. We examined deep phylogenetic relationships among the mal genes of these hyperthermophiles and their close relatives to look for evidence of shared ancestry. Results We demonstrate that the two maltose ATP binding cassette (ABC transporter operons now found in Tc. litoralis and P. furiosus (termed mal and mdx genes, respectively are not closely related to one another. The Tc. litoralis and P. furiosus mal genes are most closely related to bacterial mal genes while their respective mdx genes are archaeal. The genes of the two mal operons in Tt. maritima are not related to genes in either of these archaeal operons. They are highly similar to one another and belong to a phylogenetic lineage that includes mal genes from the enteric bacteria. A unique domain of the enteric MalF membrane spanning proteins found also in these Thermotogales MalF homologs supports their relatively close relationship with these enteric proteins. Analyses of genome sequence data from other Thermotogales species, Fervidobacterium nodosum, Thermosipho melanesiensis, Thermotoga petrophila, Thermotoga lettingae, and Thermotoga neapolitana, revealed a third apparent mal operon, absent from the published genome sequence of Tt. maritima strain MSB8. This third operon, mal3, is more closely related to the Thermococcales' bacteria-derived mal genes than are mal1 and mal2. F. nodosum, Ts. melanesiensis, and Tt. lettingae have only one of the mal1-mal2 paralogs. The mal2 operon from an unknown species of Thermotoga appears to

  14. PPAR Medicines and Human Disease: The ABCs of It All

    Directory of Open Access Journals (Sweden)

    Anthony J. Apostoli

    2012-01-01

    Full Text Available ATP-dependent binding cassette (ABC transporters are a family of transmembrane proteins that pump a variety of hydrophobic compounds across cellular and subcellular barriers and are implicated in human diseases such as cancer and atherosclerosis. Inhibition of ABC transporter activity showed promise in early preclinical studies; however, the outcomes in clinical trials with these agents have not been as encouraging. Peroxisome proliferator-activated receptors (PPARs are ligand-activated transcription factors that regulate genes involved in fat and glucose metabolism, and inflammation. Activation of PPAR signaling is also reported to regulate ABC gene expression. This suggests the potential of PPAR medicines as a novel means of controlling ABC transporter activity at the transcriptional level. This paper summarizes the advances made in understanding how PPAR medicines affect ABC transporters, and the potential implications for impacting on human diseases, in particular with respect to cancer and atherosclerosis.

  15. ATP binding and hydrolysis by Saccharomyces cerevisiae Msh2-Msh3 are differentially modulated by Mismatch and Double-strand Break Repair DNA substrates

    Science.gov (United States)

    Kumar, Charanya; Eichmiller, Robin; Wang, Bangchen; Williams, Gregory M.; Bianco, Piero R.; Surtees, Jennifer A.

    2014-01-01

    In Saccharomyces cerevisiae, Msh2-Msh3-mediated mismatch repair (MMR) recognizes and targets insertion/deletion loops for repair. Msh2-Msh3 is also required for 3′ non-homologous tail removal (3′NHTR) in double-strand break repair. In both pathways, Msh2-Msh3 binds double-strand/single-strand junctions and initiates repair in an ATP-dependent manner. However, we recently demonstrated that the two pathways have distinct requirements with respect to Msh2-Msh3 activities. We identified a set of aromatic residues in the nucleotide binding pocket (FLY motif) of Msh3 that, when mutated, disrupted MMR, but left 3′ NHTR largely intact. One of these mutations, msh3Y942A, was predicted to disrupt the nucleotide sandwich and allow altered positioning of ATP within the pocket. To develop a mechanistic understanding of the differential requirements for ATP binding and/or hydrolysis in the two pathways, we characterized Msh2-Msh3 and Msh2-msh3Y942A ATP binding and hydrolysis activities in the presence of MMR and 3′ NHTR DNA substrates. We observed distinct, substrate-dependent ATP hydrolysis and nucleotide turnover by Msh2-Msh3, indicating that the MMR and 3′ NHTR DNA substrates differentially modify the ATP binding/hydrolysis activities of Msh2-Msh3. Msh2-msh3Y942A retained the ability to bind DNA and ATP but exhibited altered ATP hydrolysis and nucleotide turnover. We propose that both ATP and structure-specific repair substrates cooperate to direct Msh2-Msh3-mediated repair and suggest an explanation for the msh3Y942A separation-of-function phenotype. PMID:24746922

  16. Structural models of zebrafish (Danio rerio NOD1 and NOD2 NACHT domains suggest differential ATP binding orientations: insights from computational modeling, docking and molecular dynamics simulations.

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    Jitendra Maharana

    Full Text Available Nucleotide-binding oligomerization domain-containing protein 1 (NOD1 and NOD2 are cytosolic pattern recognition receptors playing pivotal roles in innate immune signaling. NOD1 and NOD2 recognize bacterial peptidoglycan derivatives iE-DAP and MDP, respectively and undergoes conformational alternation and ATP-dependent self-oligomerization of NACHT domain followed by downstream signaling. Lack of structural adequacy of NACHT domain confines our understanding about the NOD-mediated signaling mechanism. Here, we predicted the structure of NACHT domain of both NOD1 and NOD2 from model organism zebrafish (Danio rerio using computational methods. Our study highlighted the differential ATP binding modes in NOD1 and NOD2. In NOD1, γ-phosphate of ATP faced toward the central nucleotide binding cavity like NLRC4, whereas in NOD2 the cavity was occupied by adenine moiety. The conserved 'Lysine' at Walker A formed hydrogen bonds (H-bonds and Aspartic acid (Walker B formed electrostatic interaction with ATP. At Sensor 1, Arg328 of NOD1 exhibited an H-bond with ATP, whereas corresponding Arg404 of NOD2 did not. 'Proline' of GxP motif (Pro386 of NOD1 and Pro464 of NOD2 interacted with adenine moiety and His511 at Sensor 2 of NOD1 interacted with γ-phosphate group of ATP. In contrast, His579 of NOD2 interacted with the adenine moiety having a relatively inverted orientation. Our findings are well supplemented with the molecular interaction of ATP with NLRC4, and consistent with mutagenesis data reported for human, which indicates evolutionary shared NOD signaling mechanism. Together, this study provides novel insights into ATP binding mechanism, and highlights the differential ATP binding modes in zebrafish NOD1 and NOD2.

  17. Hyperactivity of the Arabidopsis cryptochrome (cry1) L407F mutant is caused by a structural alteration close to the cry1 ATP-binding site.

    Science.gov (United States)

    Orth, Christian; Niemann, Nils; Hennig, Lars; Essen, Lars-Oliver; Batschauer, Alfred

    2017-08-04

    Plant cryptochromes (cry) act as UV-A/blue light receptors. The prototype, Arabidopsis thaliana cry1, regulates several light responses during the life cycle, including de-etiolation, and is also involved in regulating flowering time. The cry1 photocycle is initiated by light absorption by its FAD chromophore, which is most likely fully oxidized (FAD ox ) in the dark state and photoreduced to the neutral flavin semiquinone (FADH°) in its lit state. Cryptochromes lack the DNA-repair activity of the closely related DNA photolyases, but they retain the ability to bind nucleotides such as ATP. The previously characterized L407F mutant allele of Arabidopsis cry1 is biologically hyperactive and seems to mimic the ATP-bound state of cry1, but the reason for this phenotypic change is unclear. Here, we show that cry1 L407F can still bind ATP, has less pronounced photoreduction and formation of FADH° than wild-type cry1, and has a dark reversion rate 1.7 times lower than that of the wild type. The hyperactivity of cry1 L407F is not related to a higher FADH° occupancy of the photoreceptor but is caused by a structural alteration close to the ATP-binding site. Moreover, we show that ATP binds to cry1 in both the dark and the lit states. This binding was not affected by cry1's C-terminal extension, which is important for signal transduction. Finally, we show that a recently discovered chemical inhibitor of cry1, 3-bromo-7-nitroindazole, competes for ATP binding and thereby diminishes FADH° formation, which demonstrates that both processes are important for cry1 function. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  18. Phe(475) and Glu(446) but not Ser(445) participate in ATP-binding to the alpha-subunit of Na(+)/K(+)-ATPase

    Czech Academy of Sciences Publication Activity Database

    Kubala, Martin; Hofbauerová, Kateřina; Ettrich, Rüdiger; Kopecký ml., Vladimír; Krumscheid, R.; Plášek, J.; Teisinger, Jan; Schoner, W.; Amler, Evžen

    2002-01-01

    Roč. 297, č. 1 (2002), s. 154-159 ISSN 0006-291X R&D Projects: GA ČR GA204/01/0254; GA ČR GA204/01/1001 Grant - others:Germany(DE) WTZ CZE 00/033; Volkswagen Foundation(DE) I/74 679 Institutional research plan: CEZ:AV0Z5011922 Keywords : Na(+)/K(+)-ATPase * fluorescence spectroscopy * ATP-binding site Subject RIV: CE - Biochemistry Impact factor: 2.935, year: 2002

  19. Hepatic ABC transporters and triglyceride metabolism.

    Science.gov (United States)

    Parks, John S; Chung, Soonkyu; Shelness, Gregory S

    2012-06-01

    Elevated plasma triglyceride and reduced HDL concentrations are prominent features of metabolic syndrome and type 2 diabetes. Individuals with Tangier disease also have elevated plasma triglyceride concentrations and very low HDL, resulting from mutations in ATP-binding cassette transporter A1 (ABCA1), an integral membrane protein that facilitates nascent HDL particle assembly. Past studies attributed the inverse relationship between plasma HDL and triglyceride to intravascular lipid exchange and catabolic events. However, recent studies also suggest that hepatic signaling and lipid mobilization and secretion may explain how HDL affects plasma triglyceride concentrations. Hepatocyte-specific ABCA1 knockout mice have markedly reduced plasma HDL and a two-fold increase in triglyceride due to failure to assemble nascent HDL particles by hepatocytes, causing increased catabolism of HDL apolipoprotein A-I and increased hepatic production of triglyceride-enriched VLDL. In-vitro studies suggest that nascent HDL particles may induce signaling to decrease triglyceride secretion. Inhibition of microRNA 33 expression in nonhuman primates augments hepatic ABCA1, genes involved in fatty acid oxidation, and decreases expression of lipogenic genes, causing increased plasma HDL and decreased triglyceride levels. New evidence suggests potential mechanisms by which hepatic ABCA1-mediated nascent HDL formation regulates VLDL-triglyceride production and contributes to the inverse relationship between plasma HDL and triglyceride.

  20. Effects of in vitro exposure to ivermectin and levamisole on the expression patterns of ABC transporters in Haemonchus contortus larvae

    Directory of Open Access Journals (Sweden)

    Ali Raza

    2016-08-01

    Full Text Available This study investigated the interaction of ATP binding cassette (ABC transport proteins with ivermectin (IVM and levamisole (LEV in larvae of susceptible and resistant isolates of Haemonchus contortus in vitro by measuring transcription patterns following exposure to these anthelmintics. Furthermore, we studied the consequences of drug exposure by measuring the sensitivity of L3 to subsequent exposure to higher drug concentrations using larval migration assays. The most highly transcribed transporter genes in both susceptible and resistant L3 were pgp-9.3, abcf-1, mrp-5, abcf-2, pgp-3, and pgp-10. The resistant isolate showed significantly higher transcription of pgp-1, pgp-9.1 and pgp-9.2 compared to the susceptible isolate. Five P-gp genes and the haf-6 gene showed significantly higher transcription (up to 12.6-fold after 3 h exposure to IVM in the resistant isolate. Similarly, five P-gp genes, haf-6 and abcf-1 were transcribed at significantly higher levels (up to 10.3-fold following 3 h exposure to LEV in this isolate. On the other hand, there were no significant changes in transcriptional patterns of all transporter genes in the susceptible isolate following 3 and 6 h exposure to IVM or LEV. In contrast to these isolate-specific transcription changes, both isolates showed an increase in R-123 efflux following exposure to the drugs, suggesting that the drugs stimulated activity of existing transporter proteins in both isolates. Exposure of resistant larvae to IVM or LEV resulted, in some instances, in an increase in the proportion of the population able to migrate at the highest IVM concentrations in subsequent migration assays. The significant increase in transcription of some ABC transporter genes following 3 h exposure to both IVM and LEV in the resistant isolate only, suggests that an ability to rapidly upregulate protective pathways in response to drugs may be a component of the resistance displayed by this isolate.

  1. Involvement of the carboxyl-terminal region of the yeast peroxisomal half ABC transporter Pxa2p in its interaction with Pxa1p and in transporter function.

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    Cheng-Yi Chuang

    Full Text Available The peroxisome is a single membrane-bound organelle in eukaryotic cells involved in lipid metabolism, including β-oxidation of fatty acids. The human genetic disorder X-linked adrenoleukodystrophy (X-ALD is caused by mutations in the ABCD1 gene (encoding ALDP, a peroxisomal half ATP-binding cassette [ABC] transporter. This disease is characterized by defective peroxisomal β-oxidation and a large accumulation of very long-chain fatty acids in brain white matter, adrenal cortex, and testis. ALDP forms a homodimer proposed to be the functional transporter, whereas the peroxisomal transporter in yeast is a heterodimer comprising two half ABC transporters, Pxa1p and Pxa2p, both orthologs of human ALDP. While the carboxyl-terminal domain of ALDP is engaged in dimerization, it remains unknown whether the same region is involved in the interaction between Pxa1p and Pxa2p.Using a yeast two-hybrid assay, we found that the carboxyl-terminal region (CT of Pxa2p, but not of Pxa1p, is required for their interaction. Further analysis indicated that the central part of the CT (designated CT2 of Pxa2p was indispensable for its interaction with the carboxyl terminally truncated Pxa1_NBD. An interaction between the CT of Pxa2p and Pxa1_NBD was not detected, but could be identified in the presence of Pxa2_NBD-CT1. A single mutation of two conserved residues (aligned with X-ALD-associated mutations at the same positions in ALDP in the CT2 of the Pxa2_NBD-CT protein impaired its interaction with Pxa1_NBD or Pxa1_NBD-CT, resulting in a mutant protein that exhibited a proteinase K digestion profile different from that of the wild-type protein. Functional analysis of these mutant proteins on oleate plates indicated that they were defective in transporter function.The CT of Pxa2p is involved in its interaction with Pxa1p and in transporter function. This concept may be applied to human ALDP studies, helping to establish the pathological mechanism for CT-related X

  2. Zinc and ATP binding of the hexameric AAA-ATPase PilF from Thermus thermophilus: role in complex stability, piliation, adhesion, twitching motility, and natural transformation.

    Science.gov (United States)

    Salzer, Ralf; Herzberg, Martin; Nies, Dietrich H; Joos, Friederike; Rathmann, Barbara; Thielmann, Yvonne; Averhoff, Beate

    2014-10-31

    The traffic AAA-ATPase PilF is essential for pilus biogenesis and natural transformation of Thermus thermophilus HB27. Recently, we showed that PilF forms hexameric complexes containing six zinc atoms coordinated by conserved tetracysteine motifs. Here we report that zinc binding is essential for complex stability. However, zinc binding is neither required for pilus biogenesis nor natural transformation. A number of the mutants did not exhibit any pili during growth at 64 °C but still were transformable. This leads to the conclusion that type 4 pili and the DNA translocator are distinct systems. At lower growth temperatures (55 °C) the zinc-depleted multiple cysteine mutants were hyperpiliated but defective in pilus-mediated twitching motility. This provides evidence that zinc binding is essential for the role of PilF in pilus dynamics. Moreover, we found that zinc binding is essential for complex stability but dispensable for ATPase activity. In contrast to many polymerization ATPases from mesophilic bacteria, ATP binding is not required for PilF complex formation; however, it significantly increases complex stability. These data suggest that zinc and ATP binding increase complex stability that is important for functionality of PilF under extreme environmental conditions. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  3. Combined phylogeny and neighborhood analysis of the evolution of the ABC transporters conferring multiple drug resistance in hemiascomycete yeasts

    Directory of Open Access Journals (Sweden)

    Goffeau André

    2009-10-01

    Full Text Available Abstract Background Pleiotropic Drug Resistant transporters (PDR are members of the ATP-Binding Cassette (ABC subfamily which export antifungals and other xenobiotics in fungi and plants. This subfamily of transmembrane transporters has nine known members in Saccharomyces cerevisiae. We have analyzed the complex evolution of the pleiotropic drug resistance proteins (Pdrp subfamily where gene duplications and deletions occur independently in individual genomes. This study was carried out on 62 Pdrp from nine hemiascomycetous species, seven of which span 6 of the 14 clades of the Saccharomyces complex while the two others species, Debaryomyces hansenii and Yarrowia lipolytica, are further apart from an evolutive point of view. Results Combined phylogenetic and neighborhood analyses enabled us to identify five Pdrp clusters in the Saccharomyces complex. Three of them comprise orthologs of the Pdrp sensu stricto, Pdr5p, Pdr10p, Pdr12p, Pdr15p, Snq2p and YNR070wp. The evolutive pathway of the orthologs of Snq2 and YNR070w is particularly complex due to a tandem gene array in Eremothecium gossypii, Kluyveromyces lactis and Saccharomyces (Lachancea kluyveri. This pathway and different cases of duplications and deletions were clarified by using a neighborhood analysis based on synteny. For the two distant species, Yarrowia lipolytica and Debaryomyces hansenii, no neighborhood evidence is available for these clusters and many homologs of Pdr5 and Pdr15 are phylogenetically assigned to species-based clusters. Two other clusters comprise the orthologs of the sensu lato Pdrp, Aus1p/Pdr11p and YOL075cp respectively. The evolutionary pathway of these clusters is simpler. Nevertheless, orthologs of these genes are missing in some species. Conclusion Numerous duplications were traced among the Hemiascomycetous Pdrp studied. The role of the Whole Genome Duplication (WGD is sorted out and our analyses confirm the common ancestrality of Pdr5p and Pdr15p. A tandem

  4. Combined phylogeny and neighborhood analysis of the evolution of the ABC transporters conferring multiple drug resistance in hemiascomycete yeasts.

    Science.gov (United States)

    Seret, Marie-Line; Diffels, Julie F; Goffeau, André; Baret, Philippe V

    2009-10-01

    Pleiotropic Drug Resistant transporters (PDR) are members of the ATP-Binding Cassette (ABC) subfamily which export antifungals and other xenobiotics in fungi and plants. This subfamily of transmembrane transporters has nine known members in Saccharomyces cerevisiae. We have analyzed the complex evolution of the pleiotropic drug resistance proteins (Pdrp) subfamily where gene duplications and deletions occur independently in individual genomes. This study was carried out on 62 Pdrp from nine hemiascomycetous species, seven of which span 6 of the 14 clades of the Saccharomyces complex while the two others species, Debaryomyces hansenii and Yarrowia lipolytica, are further apart from an evolutive point of view. Combined phylogenetic and neighborhood analyses enabled us to identify five Pdrp clusters in the Saccharomyces complex. Three of them comprise orthologs of the Pdrp sensu stricto, Pdr5p, Pdr10p, Pdr12p, Pdr15p, Snq2p and YNR070wp. The evolutive pathway of the orthologs of Snq2 and YNR070w is particularly complex due to a tandem gene array in Eremothecium gossypii, Kluyveromyces lactis and Saccharomyces (Lachancea) kluyveri. This pathway and different cases of duplications and deletions were clarified by using a neighborhood analysis based on synteny. For the two distant species, Yarrowia lipolytica and Debaryomyces hansenii, no neighborhood evidence is available for these clusters and many homologs of Pdr5 and Pdr15 are phylogenetically assigned to species-based clusters. Two other clusters comprise the orthologs of the sensu lato Pdrp, Aus1p/Pdr11p and YOL075cp respectively. The evolutionary pathway of these clusters is simpler. Nevertheless, orthologs of these genes are missing in some species. Numerous duplications were traced among the Hemiascomycetous Pdrp studied. The role of the Whole Genome Duplication (WGD) is sorted out and our analyses confirm the common ancestrality of Pdr5p and Pdr15p. A tandem gene array is observed in Eremothecium gossypii. One

  5. Decipher the mechanisms of protein conformational changes induced by nucleotide binding through free-energy landscape analysis: ATP binding to Hsp70.

    Directory of Open Access Journals (Sweden)

    Adrien Nicolaï

    Full Text Available ATP regulates the function of many proteins in the cell by transducing its binding and hydrolysis energies into protein conformational changes by mechanisms which are challenging to identify at the atomic scale. Based on molecular dynamics (MD simulations, a method is proposed to analyze the structural changes induced by ATP binding to a protein by computing the effective free-energy landscape (FEL of a subset of its coordinates along its amino-acid sequence. The method is applied to characterize the mechanism by which the binding of ATP to the nucleotide-binding domain (NBD of Hsp70 propagates a signal to its substrate-binding domain (SBD. Unbiased MD simulations were performed for Hsp70-DnaK chaperone in nucleotide-free, ADP-bound and ATP-bound states. The simulations revealed that the SBD does not interact with the NBD for DnaK in its nucleotide-free and ADP-bound states whereas the docking of the SBD was found in the ATP-bound state. The docked state induced by ATP binding found in MD is an intermediate state between the initial nucleotide-free and final ATP-bound states of Hsp70. The analysis of the FEL projected along the amino-acid sequence permitted to identify a subset of 27 protein internal coordinates corresponding to a network of 91 key residues involved in the conformational change induced by ATP binding. Among the 91 residues, 26 are identified for the first time, whereas the others were shown relevant for the allosteric communication of Hsp70 s in several experiments and bioinformatics analysis. The FEL analysis revealed also the origin of the ATP-induced structural modifications of the SBD recently measured by Electron Paramagnetic Resonance. The pathway between the nucleotide-free and the intermediate state of DnaK was extracted by applying principal component analysis to the subset of internal coordinates describing the transition. The methodology proposed is general and could be applied to analyze allosteric communication in

  6. Overcoming ABC transporter-mediated multidrug resistance: Molecular mechanisms and novel therapeutic drug strategies.

    Science.gov (United States)

    Li, Wen; Zhang, Han; Assaraf, Yehuda G; Zhao, Kun; Xu, Xiaojun; Xie, Jinbing; Yang, Dong-Hua; Chen, Zhe-Sheng

    2016-07-01

    Multidrug resistance is a key determinant of cancer chemotherapy failure. One of the major causes of multidrug resistance is the enhanced efflux of drugs by membrane ABC transporters. Targeting ABC transporters projects a promising approach to eliminating or suppressing drug resistance in cancer treatment. To reveal the functional mechanisms of ABC transporters in drug resistance, extensive studies have been conducted from identifying drug binding sites to elucidating structural dynamics. In this review article, we examined the recent crystal structures of ABC proteins to depict the functionally important structural elements, such as domains, conserved motifs, and critical amino acids that are involved in ATP-binding and drug efflux. We inspected the drug-binding sites on ABC proteins and the molecular mechanisms of various substrate interactions with the drug binding pocket. While our continuous battle against drug resistance is far from over, new approaches and technologies have emerged to push forward our frontier. Most recent developments in anti-MDR strategies include P-gp inhibitors, RNA-interference, nano-medicines, and delivering combination strategies. With the advent of the 'Omics' era - genomics, epigenomics, transcriptomics, proteomics, and metabolomics - these disciplines play an important role in fighting the battle against chemoresistance by further unraveling the molecular mechanisms of drug resistance and shed light on medical therapies that specifically target MDR. Copyright © 2016 Elsevier Ltd. All rights reserved.

  7. Characterization of a novel domain ‘GATE’ in the ABC protein DrrA and its role in drug efflux by the DrrAB complex

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Han; Rahman, Sadia; Li, Wen; Fu, Guoxing; Kaur, Parjit, E-mail: pkaur@gsu.edu

    2015-03-27

    A novel domain, GATE (Glycine-loop And Transducer Element), is identified in the ABC protein DrrA. This domain shows sequence and structural conservation among close homologs of DrrA as well as distantly-related ABC proteins. Among the highly conserved residues in this domain are three glycines, G215, G221 and G231, of which G215 was found to be critical for stable expression of the DrrAB complex. Other conserved residues, including E201, G221, K227 and G231, were found to be critical for the catalytic and transport functions of the DrrAB transporter. Structural analysis of both the previously published crystal structure of the DrrA homolog MalK and the modeled structure of DrrA showed that G215 makes close contacts with residues in and around the Walker A motif, suggesting that these interactions may be critical for maintaining the integrity of the ATP binding pocket as well as the complex. It is also shown that G215A or K227R mutation diminishes some of the atomic interactions essential for ATP catalysis and overall transport function. Therefore, based on both the biochemical and structural analyses, it is proposed that the GATE domain, located outside of the previously identified ATP binding and hydrolysis motifs, is an additional element involved in ATP catalysis. - Highlights: • A novel domain ‘GATE’ is identified in the ABC protein DrrA. • GATE shows high sequence and structural conservation among diverse ABC proteins. • GATE is located outside of the previously studied ATP binding and hydrolysis motifs. • Conserved GATE residues are critical for stability of DrrAB and for ATP catalysis.

  8. Only one ATP-binding DnaX subunit is required for initiation complex formation by the Escherichia coli DNA polymerase III holoenzyme.

    Science.gov (United States)

    Wieczorek, Anna; Downey, Christopher D; Dallmann, H Garry; McHenry, Charles S

    2010-09-17

    The DnaX complex (DnaX(3)δδ'χ psi) within the Escherichia coli DNA polymerase III holoenzyme serves to load the dimeric sliding clamp processivity factor, β(2), onto DNA. The complex contains three DnaX subunits, which occur in two forms: τ and the shorter γ, produced by translational frameshifting. Ten forms of E. coli DnaX complex containing all possible combinations of wild-type or a Walker A motif K51E variant τ or γ have been reconstituted and rigorously purified. DnaX complexes containing three DnaX K51E subunits do not bind ATP. Comparison of their ability to support formation of initiation complexes, as measured by processive replication by the DNA polymerase III holoenzyme, indicates a minimal requirement for one ATP-binding DnaX subunit. DnaX complexes containing two mutant DnaX subunits support DNA synthesis at about two-thirds the level of their wild-type counterparts. β(2) binding (determined functionally) is diminished 12-30-fold for DnaX complexes containing two K51E subunits, suggesting that multiple ATPs must be bound to place the DnaX complex into a conformation with maximal affinity for β(2). DNA synthesis activity can be restored by increased concentrations of β(2). In contrast, severe defects in ATP hydrolysis are observed upon introduction of a single K51E DnaX subunit. Thus, ATP binding, hydrolysis, and the ability to form initiation complexes are not tightly coupled. These results suggest that although ATP hydrolysis likely enhances β(2) loading, it is not absolutely required in a mechanistic sense for formation of functional initiation complexes.

  9. Computer modelling reveals new conformers of the ATP binding loop of Na+/K+-ATPase involved in the transphosphorylation process of the sodium pump.

    Science.gov (United States)

    Tejral, Gracian; Sopko, Bruno; Necas, Alois; Schoner, Wilhelm; Amler, Evzen

    2017-01-01

    Hydrolysis of ATP by Na + /K + -ATPase, a P-Type ATPase, catalyzing active Na + and K + transport through cellular membranes leads transiently to a phosphorylation of its catalytical α -subunit. Surprisingly, three-dimensional molecular structure analysis of P-type ATPases reveals that binding of ATP to the N-domain connected by a hinge to the P-domain is much too far away from the Asp 369 to allow the transfer of ATP's terminal phosphate to its aspartyl-phosphorylation site. In order to get information for how the transfer of the γ -phosphate group of ATP to the Asp 369 is achieved, analogous molecular modeling of the M 4 -M 5 loop of ATPase was performed using the crystal data of Na + /K + -ATPase of different species. Analogous molecular modeling of the cytoplasmic loop between Thr 338 and Ile 760 of the α 2 -subunit of Na + /K + -ATPase and the analysis of distances between the ATP binding site and phosphorylation site revealed the existence of two ATP binding sites in the open conformation; the first one close to Phe 475 in the N-domain, the other one close to Asp 369 in the P-domain. However, binding of Mg 2+ •ATP to any of these sites in the "open conformation" may not lead to phosphorylation of Asp 369 . Additional conformations of the cytoplasmic loop were found wobbling between "open conformation"  "semi-open conformation  "closed conformation" in the absence of 2Mg 2+ •ATP. The cytoplasmic loop's conformational change to the "semi-open conformation"-characterized by a hydrogen bond between Arg 543 and Asp 611 -triggers by binding of 2Mg 2+ •ATP to a single ATP site and conversion to the "closed conformation" the phosphorylation of Asp 369 in the P-domain, and hence the start of Na + /K + -activated ATP hydrolysis.

  10. ABC Kinga kauplused = ABC King shoe stores

    Index Scriptorium Estoniae

    2011-01-01

    Tallinnas Kristiine keskuses, Tartu Kaubamajas ja Pärnus Port Artur 2 asuvate ABC Kinga kaupluste sisekujundusest. Sisearhitekid Andres Labi ja Janno Roos (Ruumilabor OÜ), loetletud nende ühiselt tehtud töid

  11. Bacterial multidrug resistance mediated by a homologue of the human multidrug transporter P-glycoprotein

    NARCIS (Netherlands)

    Konings, WN; Poelarends, GJ

    2002-01-01

    Most ATP-binding cassette (ABC) multidrug transporters known to date are of eukaryotic origin, such as the P-glycoproteins (Pgps) and multidrug resistance-associated proteins (MRPs). Only one well-characterized ABC multidrug transporter, LmrA, is of bacterial origin. On the basis of its structural

  12. Effect of (mixtures of) flavonoids on the in vitro and in vivo bioavailability of 2-mino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) : a biologically based modelling approach

    NARCIS (Netherlands)

    Schutte, M.E.

    2007-01-01

    The transport of food ingredients across the intestinal epithelium is an important factor determining the absorption upon oral intake. Uptake of compounds in the intestine may be influenced by transport proteins such as the ATP binding cassette transporters (ABC transporters). ABC transporters have

  13. Inactivation of the peroxisomal ABCD2 transporter in the mouse leads to late-onset ataxia involving mitochondria, Golgi and endoplasmic reticulum damage

    NARCIS (Netherlands)

    Ferrer, Isidre; Kapfhammer, Josef P.; Hindelang, Colette; Kemp, Stephan; Troffer-Charlier, Nathalie; Broccoli, Vania; Callyzot, Noëlle; Mooyer, Petra; Selhorst, Jacqueline; Vreken, Peter; Wanders, Ronald J. A.; Mandel, Jean Louis; Pujol, Aurora

    2005-01-01

    ATP-binding cassette (ABC) transporters facilitate unidirectional translocation of chemically diverse substances, ranging from peptides to lipids, across cell or organelle membranes. In peroxisomes, a subfamily of four ABC transporters (ABCD1 to ABCD4) has been related to fatty acid transport,

  14. ABCA Transporter Gene Expression and Poor Outcome in Epithelial Ovarian Cancer

    DEFF Research Database (Denmark)

    Hedditch, Ellen L; Gao, Bo; Russell, Amanda J

    2014-01-01

    BACKGROUND: ATP-binding cassette (ABC) transporters play various roles in cancer biology and drug resistance, but their association with outcomes in serous epithelial ovarian cancer (EOC) is unknown. METHODS: The relationship between clinical outcomes and ABC transporter gene expression in two in...

  15. Enhancing the ABC Cross

    OpenAIRE

    Euske, K.J.; Vercio, Alan

    2007-01-01

    The purpose of the ABC Cross was to portray both a cost and process view of an organization as simply as possible. Unfortunately, the model’s simplified form does not capture the real value of activity-based costing (ABC) for cost accounting that emerged in the mid-1980s. Here we present several ABC models that can help functional and process managers make better decisions.

  16. The Absence of the Transcription Factor Yrr1p, Identified from Comparative Genome Profiling, Increased Vanillin Tolerance Due to Enhancements of ABC Transporters Expressing, rRNA Processing and Ribosome Biogenesis in Saccharomyces cerevisiae.

    Science.gov (United States)

    Wang, Xinning; Liang, Zhenzhen; Hou, Jin; Shen, Yu; Bao, Xiaoming

    2017-01-01

    Enhancing the tolerance of Saccharomyces cerevisiae to inhibitors derived from lignocellulose is conducive to producing biofuel and chemicals using abundant lignocellulosic materials. Vanillin is a major type of phenolic inhibitor in lignocellulose hydrolysates for S. cerevisiae . In the present work, the factors beneficial to vanillin resistance in yeast were identified from the vanillin-resistant strain EMV-8, which was derived from strain NAN-27 by adaptive evolution. We found 450 SNPs and 44 genes with InDels in the vanillin-tolerant strain EMV-8 by comparing the genome sequences of EMV-8 and NAN-27. To investigate the effects of InDels, InDels were deleted in BY4741, respectively. We demonstrated that the deletion of YRR1 improved vanillin tolerance of strain. In the presence of 6 mM vanillin, deleting YRR1 increase the maximum specific growth rate and the vanillin consumption rate by 142 and 51%, respectively. The subsequent transcriptome analysis revealed that deleting YRR1 resulted in changed expression of over 200 genes in the presence of 5 mM vanillin. The most marked changes were the significant up-regulation of the dehydrogenase ADH7 , several ATP-binding cassette (ABC) transporters, and dozens of genes involved in ribosome biogenesis and rRNA processing. Coincidently, the crude enzyme solution of BY4741( yrr1 Δ) exhibited higher NADPH-dependent vanillin reduction activity than control. In addition, overexpressing the ABC transporter genes PDR5, YOR1 , and SNQ2 , as well as the RNA helicase gene DBP2 , increased the vanillin tolerance of strain. Interestingly, unlike the marked changes we mentioned above, under vanillin-free conditions, there are only limited transcriptional differences between wildtype and yrr1 Δ. This indicated that vanillin might act as an effector in Yrr1p-related regulatory processes. The new findings of the relationship between YRR1 and vanillin tolerance, as well as the contribution of rRNA processing and ribosome biogenesis to

  17. Computer modelling reveals new conformers of the ATP binding loop of Na+/K+-ATPase involved in the transphosphorylation process of the sodium pump

    Directory of Open Access Journals (Sweden)

    Gracian Tejral

    2017-03-01

    Full Text Available Hydrolysis of ATP by Na+/K+-ATPase, a P-Type ATPase, catalyzing active Na+ and K+ transport through cellular membranes leads transiently to a phosphorylation of its catalytical α-subunit. Surprisingly, three-dimensional molecular structure analysis of P-type ATPases reveals that binding of ATP to the N-domain connected by a hinge to the P-domain is much too far away from the Asp369 to allow the transfer of ATP’s terminal phosphate to its aspartyl-phosphorylation site. In order to get information for how the transfer of the γ-phosphate group of ATP to the Asp369 is achieved, analogous molecular modeling of the M4–M5 loop of ATPase was performed using the crystal data of Na+/K+-ATPase of different species. Analogous molecular modeling of the cytoplasmic loop between Thr338 and Ile760 of the α2-subunit of Na+/K+-ATPase and the analysis of distances between the ATP binding site and phosphorylation site revealed the existence of two ATP binding sites in the open conformation; the first one close to Phe475 in the N-domain, the other one close to Asp369 in the P-domain. However, binding of Mg2+•ATP to any of these sites in the “open conformation” may not lead to phosphorylation of Asp369. Additional conformations of the cytoplasmic loop were found wobbling between “open conformation”  “semi-open conformation  “closed conformation” in the absence of 2Mg2+•ATP. The cytoplasmic loop’s conformational change to the “semi-open conformation”—characterized by a hydrogen bond between Arg543 and Asp611—triggers by binding of 2Mg2+•ATP to a single ATP site and conversion to the “closed conformation” the phosphorylation of Asp369 in the P-domain, and hence the start of Na+/K+-activated ATP hydrolysis.

  18. The ABC and AUSSAT.

    Science.gov (United States)

    McGarritty, Ian

    1985-01-01

    Discusses the Australian Broadcasting Corporation's (ABC) utilization of the AUSSAT telecommunications satellite to extend its television and radio transmission range to reach remote Australian audiences; the satellite's program gathering and interchange capabilities; and ABC's generation of other benefits to offset cost of satellite services.…

  19. Rad51 ATP binding but not hydrolysis is required to recruit Rad10 in synthesis-dependent strand annealing sites inS. cerevisiae.

    Science.gov (United States)

    Karlin, Justin; Fischhaber, Paula L

    2013-06-01

    Several modes of eukaryotic of DNA double strand break repair (DSBR) depend on synapsis of complementary DNA. The Rad51 ATPase, the S. cerevisiae homolog of E. coli RecA, plays a key role in this process by catalyzing homology searching and strand exchange between an invading DNA strand and a repair template (e.g. sister chromatid or homologous chromosome). Synthesis dependent strand annealing (SDSA), a mode of DSBR, requires Rad51. Another repair enzyme, the Rad1-Rad10 endonuclease, acts in the final stages of SDSA, hydrolyzing 3' overhanging single-stranded DNA. Here we show in vivo by fluorescence microscopy that the ATP binding function of yeast Rad51 is required to recruit Rad10 SDSA sites indicating that Rad51 pre-synaptic filament formation must occur prior to the recruitment of Rad1-Rad10. Our data also show that Rad51 ATPase activity, an important step in Rad51 filament disassembly, is not absolutely required in order to recruit Rad1-Rad10 to DSB sites.

  20. A putative amino acid ABC transporter substrate-binding protein, NMB1612, from Neisseria meningitidis, induces murine bactericidal antibodies against meningococci expressing heterologous NMB1612 proteins.

    Science.gov (United States)

    Hung, Miao-Chiu; Humbert, María Victoria; Laver, Jay R; Phillips, Renee; Heckels, John E; Christodoulides, Myron

    2015-08-26

    The nmb1612 (NEIS1533) gene encoding the ~27-kDa putative amino acid ATP-binding cassette (ABC) transporter, periplasmic substrate-binding protein from Neisseria meningitidis serogroup B (MenB) strain MC58 was cloned and expressed in Escherichia coli, and the purified recombinant (r)NMB1612 was used for animal immunization studies. Immunization of mice with rNMB1612 adsorbed to Al(OH)3 and in liposomes with and without MPLA, induced antiserum with bactericidal activity in an assay using baby rabbit complement, against the homologous strain MC58 (encoding protein representative of Allele 62) and killed heterologous strains encoding proteins of three other alleles (representative of Alleles 1, 64 and 68), with similar SBA titres. However, strain MC58 was not killed (titre bactericidal assay (hSBA) using anti-rNMB1612 sera, although another strain (MC168) expressing the same protein was killed (median titres of 16-64 in the hSBA). Analysis of the NMB1612 amino acid sequences from 4351 meningococcal strains in the pubmlst.org/Neisseria database and a collection of 13 isolates from colonized individuals and from patients, showed that antibodies raised against rNMB1612 could potentially kill at least 72% of the MenB strains in the complete sequence database. For MenB disease occurring specifically in the UK from 2013 to 2015, >91% of the isolates causing disease in this recent period expressed NMB1612 protein encoded by Allele 1 and could be potentially killed by sera raised to the recombinant antigen in the current study. The NMB1612 protein was surface-accessible and expressed by different meningococcal strains. In summary, the properties of (i) NMB1612 protein conservation and expression, (ii) limited amino acid sequence variation between proteins encoded by different alleles, and (iii) the ability of a recombinant protein to induce cross-strain bactericidal antibodies, would all suggest a promising antigen for consideration for inclusion in new meningococcal vaccines

  1. Polymorphisms in ATP-binding cassette transporter genes and interaction with diet and life style factors in relation to colorectal cancer in a Danish prospective case-cohort study

    DEFF Research Database (Denmark)

    Kopp, Tine Iskov; Andersen, Vibeke; Tjonneland, Anne

    2015-01-01

    to assess whether polymorphisms in ABCB1, ABCC2 and ABCG2 were associated with risk of colorectal cancer (CRC) and to investigate gene-environment (dietary factors, smoking and use of non-steroidal anti-inflammatory drugs) and gene-gene interactions between previously studied polymorphisms in IL1B and IL10...

  2. [Effect of transcription activity regulated by VNTR-ZNF and -14C/T variants in the promoter region of ATP-binding cassette transporter 1 in HepG2 cells].

    Science.gov (United States)

    Gao, Shenxia; Zhao, Lili; Zhang, Ying; Mao, Yongmin

    2016-10-01

    To explore the effect of VNTR-ZNF and -14C/T variants of the promoter region of the ABCA1 gene on the transcription activity of genes in vitro. The recombinants were constructed by ligating DNA fragment containing VNTR-ZNF ACCCC inserted/deleted allele with or without -14C/T substitution fragments with a PGL2-basic vector containing luciferase reporter gene. The recombinants were then transfected into HepG2 cells using the cationic lipid method. After 48 h, transfected cells were collected and used to detect the luciferase activity. Luciferase activity of PGL2-ZNF-ACCCCDel was greater than that of PGL2-ZNF-ACCCCIns. Luciferase activity of PGL2-ZNFDel-14C was greater than that of PGL2-ZNFDel-14T, PGL2-ZNFIns-14C, PGL2-ZNFIns-14T. Compared with the insertion type, the ACCCC-deleted type of VNTR-ZNF can significantly enhance the transcription activity of ABCA1. And co-transfection of -14 C allele can further enhance this activity.

  3. Rapportering med ABC modellen

    DEFF Research Database (Denmark)

    Wiese, Lars Ole

    2006-01-01

    I denne artikel diskuteres Activity Based Costing (ABC) som rapporterings-teknik for kunde profitabilitet. Der argumenteres for, at indtægtserhvervel-sen inkl. dens direkte omkostningsforbrug bør adskilles fra omkostnings-beskrivelsen til produktfremstilling. Dvs. der gennemføres en aktivitetsba......-seret rapportering for indtægter og en ABC model for omkostningsforbrug til produktfremstilling. Modellen er tilført logiske beslutningsobjekter (klas-sifikationer), som er opbygget niveaubaseret. Samtidigt føres belæg for, at ABC modellen bør baseres på logiske fordelinger til kalkulationsobjektet i stedet...

  4. Thermodynamics of ABC transporters

    Directory of Open Access Journals (Sweden)

    Xuejun C. Zhang

    2015-09-01

    Full Text Available ABSTRACT ABC transporters form the largest of all transporter families, and their structural study has made tremendous progress over recent years. However, despite such advances, the precise mechanisms that determine the energy-coupling between ATP hydrolysis and the conformational changes following substrate binding remain to be elucidated. Here, we present our thermodynamic analysis for both ABC importers and exporters, and introduce the two new concepts of differential-binding energy and elastic conformational energy into the discussion. We hope that the structural analysis of ABC transporters will henceforth take thermodynamic aspects of transport mechanisms into account as well.

  5. Iowa ABC connections.

    Science.gov (United States)

    2015-06-01

    For several years the Iowa Department of Transportation (DOT), Iowa State University, the Federal Highway Administration, : and several Iowa counties have been working to develop accelerated bridge construction (ABC) concepts, details, and processes....

  6. Structure and function of the universal stress protein TeaD and its role in regulating the ectoine transporter TeaABC of Halomonas elongata DSM 2581(T).

    Science.gov (United States)

    Schweikhard, Eva S; Kuhlmann, Sonja I; Kunte, Hans-Jörg; Grammann, Katrin; Ziegler, Christine M

    2010-03-16

    The halophilic bacterium Halomonas elongata takes up the compatible solute ectoine via the osmoregulated TRAP transporter TeaABC. A fourth orf (teaD) is located adjacent to the teaABC locus that encodes a putative universal stress protein (USP). By RT-PCR experiments we proved a cotranscription of teaD along with teaABC. Deletion of teaD resulted in an enhanced uptake for ectoine by the transporter TeaABC and hence a negative activity regulation of TeaABC by TeaD. A transcriptional regulation via DNA binding could be excluded. ATP binding to native TeaD was shown by HPLC, and the crystal structure of TeaD was solved in complex with ATP to a resolution of 1.9 A by molecular replacement. TeaD forms a dimer-dimer complex with one ATP molecule bound to each monomer, which has a Rossmann-like alpha/beta overall fold. Our results reveal an ATP-dependent oligomerization of TeaD, which might have a functional role in the regulatory mechanism of TeaD. USP-encoding orfs, which are located adjacent to genes encoding for TeaABC homologues, could be identified in several other organisms, and their physiological role in balancing the internal cellular ectoine pool is discussed.

  7. Effect of Walker A mutation (K86M) on oligomerization and surface targeting of the multidrug resistance transporter ABCG2

    DEFF Research Database (Denmark)

    Henriksen, Ulla Birk; Gether, Ulrik; Litman, Thomas

    2005-01-01

    of the nucleotide binding domain (NBD) known to be critical for ATP binding and/or hydrolysis in ABC transporters. The mutant (ABCG2-K86M) was inactive as expected but was expressed at similar levels as the wild-type (wt) protein. The mutation did not affect the predicted oligomerization properties......The ATP binding cassette (ABC) half-transporter ABCG2 (MXR/BCRP/ABCP) is associated with mitoxantrone resistance accompanied by cross-resistance to a broad spectrum of cytotoxic drugs. Here we investigate the functional consequences of mutating a highly conserved lysine in the Walker A motif...... of the NBDs in assisting proper surface targeting of ABC transporters....

  8. ATP binding and cross-bridge detachment steps during full Ca²⁺ activation: comparison of myofibril and muscle fibre mechanics by sinusoidal analysis.

    Science.gov (United States)

    Iorga, Bogdan; Wang, Li; Stehle, Robert; Pfitzer, Gabriele; Kawai, Masataka

    2012-07-15

    Single myofibrils 50–60 μm length and 2–3 μm diameter were isolated from rabbit psoas muscle fibres, and cross-bridge kinetics were studied by small perturbations of the length (∼0.2%) over a range of 15 frequencies (1–250 Hz). The experiments were performed at 15◦C in the presence of 0.05–10 mM MgATP, 8mM phosphate (Pi), 200 mM ionic strength with KAc (acetate), pCa 4.35–4.65, and pH 7.0. Two exponential processes, B and C, were resolved in tension transients. Their apparent rate constants (2πb and 2πc) increased as the [MgATP] was raised from 0.05 mM to 1mM, and then reached saturation at [MgATP] ≥ 1. Given that these rate constants were similar (c/b ∼1.7) at [Pi] ≥ 4 mM, they were combined to achieve an accurate estimate of the kinetic constants: their sum and product were analysed as functions of [MgATP]. These analyses yielded K1 =2.91 ± 0.31 mM −1, k2 =288 ± 36 s−1, and k−2 =10 ± 21 s−1 (±95% confidence limit, n =13 preparations), based on the cross-bridge model: AM+ATP ↔ (step 1) AM.ATP ↔ (step 2) A+M.ATP, where K1 is the ATP association constant (step 1), k2 is the rate constant of the cross-bridge detachment (step 2), and k−2 is the rate constant of its reversal step. These kinetic constants are respectively comparable to those observed in single fibres from rabbit psoas (K1 =2.35 ± 0.31 mM −1, k2 =243 ± 22 s−1, and k−2 =6 ± 14 s−1; n =8 preparations) when analysed by the same methods and under the same experimental conditions. These values are respectively not significantly different from those obtained in myofibrils, indicating that the same kinetic constants can be deduced from myofibril and muscle fibre studies, in terms of ATP binding and cross-bridge detachments steps. The fact that K1 in myofibrils is 1.2 times that in fibres (P≈0.05) may be explained by a small concentration gradient of ATP, ADP and/or Pi in single fibres.

  9. Multixenobiotic resistance efflux activity in Daphnia magna and Lumbriculus variegatus

    OpenAIRE

    Vehniäinen, Eeva-Riikka; Kukkonen, Jussi

    2015-01-01

    Multixenobiotic resistance is a phenomenon in which ATP-binding cassette (ABC) family proteins transfer harmful compounds out of cells. Daphnia magna and Lumbriculus variegatus are model species in aquatic ecotoxicology, but the presence and activity of ABC proteins have not been well described in these species. The aim of this work was to study the presence, activity, and inhibition of ABC transport proteins in D. magna and L. variegatus. The presence of abcb1 and abcc transcripts in 8–9-day...

  10. Plant ABC Transporters

    Science.gov (United States)

    Kang, Joohyun; Park, Jiyoung; Choi, Hyunju; Burla, Bo; Kretzschmar, Tobias; Lee, Youngsook; Martinoia, Enrico

    2011-01-01

    ABC transporters constitute one of the largest protein families found in all living organisms. ABC transporters are driven by ATP hydrolysis and can act as exporters as well as importers. The plant genome encodes for more than 100 ABC transporters, largely exceeding that of other organisms. In Arabidopsis, only 22 out of 130 have been functionally analyzed. They are localized in most membranes of a plant cell such as the plasma membrane, the tonoplast, chloroplasts, mitochondria and peroxisomes and fulfill a multitude of functions. Originally identified as transporters involved in detoxification processes, they have later been shown to be required for organ growth, plant nutrition, plant development, response to abiotic stresses, pathogen resistance and the interaction of the plant with its environment. To fulfill these roles they exhibit different substrate specifies by e.g. depositing surface lipids, accumulating phytate in seeds, and transporting the phytohormones auxin and abscisic acid. The aim of this review is to give an insight into the functions of plant ABC transporters and to show their importance for plant development and survival. PMID:22303277

  11. ABC's of Being Smart

    Science.gov (United States)

    Foster, Joanne

    2011-01-01

    Determining what giftedness is all about means focusing on many aspects of the individual. In this paper, the author focuses on letter D of the ABC's of being smart. She starts with specifics about giftedness (details), and then moves on to some ways of thinking (dispositions).

  12. [Research progress in chondroitinase ABC].

    Science.gov (United States)

    Li, Ye; Chen, Zhenya; Yuan, Qipeng

    2015-05-01

    As the components of proteoglycans, glycosaminoglycans (GAGs) are linear polysaccharides consisting of hexose and uronic acid units linked by β-1,3-glycosidic bond. GAGs mainly distribute in extracellular matrix and on cell surfaces. They guide many biological processes, such as proliferation of cells, transmission of signals and mediation of inflammation. Because of their large molecular weights, GAGs have limited biological functions in vitro. However, the appearance of chondroitinase ABC (ChSase ABC), which can lyse polysaccharides, solves the difficulties. Based on our work, we summarized the classification and the crystal structure of ChSase ABC, as well as other recent research progress on ChSase ABCs. The separation and purification methods of ChSase ABC and construction of engineering bacteria are illustrated. The stability and immobilization are also analyzed by taking account of the characterization of ChSase ABC. Finally, problems and future prospect of the ChSase ABC study are summarized.

  13. Mutational analysis of the binding affinity and transport activity for N-acetylglucosamine of the novel ABC transporter Ngc in the chitin-degrader Streptomyces olivaceoviridis.

    Science.gov (United States)

    Saito, A; Schrempf, H

    2004-06-01

    The highly differentiated bacterium Streptomyces olivaceoviridis efficiently hydrolyses chitin, a highly abundant natural polysaccharide, to low molecular weight products including N-acetylglucosamine (NAG) and N,N' -diacetylchitobiose (chitobiose). NAG is taken up by a PTS (phosphoenolpyruvate-dependent phosphotransferase system) which includes the PtsC2 protein, and via the ABC (ATP-binding cassette) transporter Ngc, which itself includes the substrate-binding protein NgcE. This is at present the only ABC transporter which is known to mediate specific uptake of NAG (K(m) 0.48 microM, V(max) 1.3 nmol/min/mg dry weight) and is competitively inhibited by chitobiose (K(i) 0.68 microM). The latter finding suggests that the Ngc system transports both NAG and chitobiose efficiently. To identify amino acid residues required for the function of NgcE, either the wild-type or one of several mutant forms of the ngcE gene was introduced into the strain S. olivaceoviridis DeltaNgcE/DeltaPtsC1/DeltaPtsC2, which lacks both functional transport systems for NAG, and chromosomal recombinants were selected. Based on the in vivo transport parameters of the recombinants, and the in vitro binding characteristics of the corresponding purified proteins, the following conclusions can be drawn. (1) Replacement of the C-terminally located residue Y396 by A (Y396A) has little effect on ligand-binding or transport parameters. The W395A mutation also induced little change in the substrate affinity in vitro, but it led in vivo to a marked increase (11 fold) in K(m), and enhanced V(max) (by 1.5 fold). (2) The amino acids Y201 and W280 both contribute (51% and 38%) to the ligand-binding capacity of NgcE. They are both very important for the in vivo function of the complete transport apparatus; strains expressing either Y201A or W280A show drastically (100 or 150 times) enhanced K(m) values. (3) The concomitant presence of either Y200 and W280 or Y201 and W280 is essential for the function of Ngc

  14. The ITER divertor cassette project meeting

    International Nuclear Information System (INIS)

    Merola, M.; Riccardi, B.; Tivey, R.

    1999-01-01

    The Divertor Cassette Project topical meeting was held on May 26-28, 1999 at the ENEA Brasimone Research Centre in Camugnano (Bologna), Italy. Specialists from all the four Parties and the JCT participated in the meeting. It was concluded that the Divertor Cassette Project has significantly contributed to solving a large part of the critical issues of the ITER divertor design

  15. Protein (Cyanobacteria): 652400470 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)

    Full Text Available rug ABC transporter ATP-binding protein Planktothrix prolifica MNWAIEVKDSASMSSLNPVVATQNLGKFYRTGFWMNQKIESLKSC...QMRQYSKGMLQRVGMAQALINNPEVVFLDEPMSGLDPMGRYQIREIILSLKAQNKTVFFNSHVLSDVEKICDRIAILAEGE

  16. Genome-wide identification, functional analysis and expression ...

    African Journals Online (AJOL)

    The plant pleiotropic drug resistance (PDR) family of ATP-binding cassette (ABC) transporters has comprehensively been researched in relation to transport of antifungal agents and resistant pathogens. In our study, analyses of the whole family of PDR genes present in the potato genome were provided. This analysis ...

  17. An Arabidopsis lipid flippase is required for timely recruitment of defenses to the host-pathogen interface at the plant cell surface

    Science.gov (United States)

    Deposition of cell wall-reinforcing papillae is an integral component of the plant immune response. The Arabidopsis PENETRATION 3 (PEN3) ATP binding cassette (ABC) transporter plays a role in defense against numerous pathogens and is recruited to sites of pathogen detection where it accumulates with...

  18. Lack of ABCG2 shortens latency of BRCA1-deficient mammary tumors and this is not affected by genistein or resveratrol

    NARCIS (Netherlands)

    Zander, Serge A. L.; Kersbergen, Ariena; Sol, Wendy; Gonggrijp, Maaike; van de Wetering, Koen; Jonkers, Jos; Borst, Piet; Rottenberg, Sven

    2012-01-01

    In addition to their role in drug resistance, the ATP-binding cassette (ABC) transporters ABCG2 and ABCB1 have been suggested to protect cells from a broad range of substances that may foster tumorigenesis. Phytoestrogens or their metabolites are substrates of these transporters and the influence of

  19. Abcg5/Abcg8-independent pathways contribute to hepatobiliary cholesterol secretion in mice

    NARCIS (Netherlands)

    Plosch, Torsten; van der Veen, Jelske N.; Havinga, Rick; Huijkman, Nicolette C. A.; Bloks, Vincent W.; Kuipers, Folkert

    The ATP-binding cassette (ABC) half-transporters ABCG5 and ABCG8 heterodimerize into a functional complex that mediates the secretion of plant sterols and cholesterol by hepatocytes into bile and their apical efflux from enterocytes. We addressed the putative rate-controlling role of Abcg5/Abcg8 in

  20. Genomewide analysis of ABCBs with a focus on ABCB1 and ...

    Indian Academy of Sciences (India)

    Abstract. The B subfamily of ATP-binding cassette (ABC) proteins (ABCB) plays a vital role in auxin efflux. However, no systematic study has been done in apple. In this study, we performed genomewide identification and expression analyses of the ABCB family in Malus domestica for the first time. We identified a total of 25 ...

  1. Differential substrate specificities of human ABCD1 and ABCD2 in peroxisomal fatty acid β-oxidation

    NARCIS (Netherlands)

    van Roermund, Carlo W. T.; Visser, Wouter F.; Ijlst, Lodewijk; Waterham, Hans R.; Wanders, Ronald J. A.

    2011-01-01

    The gene mutated in X-linked adrenoleukodystrophy (X-ALD) codes for the HsABCD1 protein, also named ALDP, which is a member of the superfamily of ATP-binding cassette (ABC) transporters and required for fatty acid transport across the peroxisomal membrane. Although a defective HsABCD1 results in the

  2. Journal of Genetics | Indian Academy of Sciences

    Indian Academy of Sciences (India)

    Keywords. ABCB; auxin efflux; apple; dwarf; expression pattern. Abstract. The B subfamily of ATP-binding cassette (ABC) proteins (ABCB) plays a vital role in auxin efflux. However, no systematic study has been done in apple. In this study, we performed genomewide identification and expression analyses of the ABCB ...

  3. Genetics Home Reference: Tangier disease

    Science.gov (United States)

    ... Maxfield FR, Tabas I. Role of cholesterol and lipid organization in disease. Nature. 2005 Dec 1;438(7068):612-21. ... Hubácek JA. ATP-binding cassette (ABC) transporters in human metabolism and diseases. Physiol Res. 2004;53(3):235-43. Review. ...

  4. A sensor for intracellular ionic strength

    NARCIS (Netherlands)

    Biemans-Oldehinkel, Esther; Mahmood, Nik A.B.N.; Poolman, Bert

    2006-01-01

    Cystathionine-β-synthase (CBS) domains are found in >4,000 proteins in species from all kingdoms of life, yet their functions are largely unknown. Tandem CBS domains are associated with membrane transport proteins, most notably members of the ATP-binding cassette (ABC) superfamily; voltage-gated

  5. Diversity of transport mechanisms: common structural principles

    NARCIS (Netherlands)

    Driessen, A.J.M.; Rosen, B.P.; Konings, W.N

    2000-01-01

    Traditionally, prokaryotic solute transport systems are classified into major groups based on the energetic requirement of the transport process. These include the secondary transporters that are driven by a proton or sodium motive force, and the ATP-binding cassette (ABC) primary transporters,

  6. Inhibition of multixenobiotic resistance transporters (MXR) by silver nanoparticles and ions in vitro and in Daphnia magna

    NARCIS (Netherlands)

    Georgantzopoulou, Anastasia; Cambier, Sébastien; Serchi, Tommaso; Kruszewski, Marcin; Balachandran, Yekkuni L.; Grysan, Patrick; Audinot, Jean Nicolas; Ziebel, Johanna; Guignard, Cédric; Gutleb, Arno C.; Murk, A.J.

    2016-01-01

    The P-glycoprotein (P-gp, ABCB1) and multidrug resistance associated protein 1 (MRP1), important members of the ABC (ATP-binding cassette) transporters, protect cells and organisms via efflux of xenobiotics and are responsible for the phenomenon of multidrug or multixenobiotic resistance (MXR).

  7. ABC+SCM=Sant?

    OpenAIRE

    Dahl, Jonas; Porelius, Jesper

    2006-01-01

    Background: Companies of today commonly search to gain competitive advantages throughout different forms of co-operation, one of which is referred to as Supply Chain Management. Although little research has been assigned to the topic of how to manage and control this type of relation, lately a growing number of academics has been arguing that ABC is an appropriate mean of controlling this type of relationships. Purpose: The purpose of this thesis is to investigate to what degree the ongoing d...

  8. Design of SMART steam generator cassette

    International Nuclear Information System (INIS)

    Kim, Y. W.; Kim, J. I.; Jang, M. H.

    2001-01-01

    Basic design development for the steam generator to be installed in the integral reactor SMART has been performed. Optimization of the steam generator shape, determination of the basic dimension and confirmation of the structural strength have been carried out. Individual steam generator cassette can be replaced in the optimized design concept of steam generator. Shape design of the steam generator cassette has been done on the computer based on 3-D CAE strategy. The structural integrity of the developed steam generator was investigated by performing the dynamic analysis for the steam generator cassette, flow induced vibration analysis for the tube bundle, and the thermo-mechanical analysis for the module header and tube. As for the manufacturing of steam generator, the numerical and the experimental simulation have been carried to control the amount of spring back and to eliminate residual stress. SMART steam generator cassette was developed by a sequential research of the aforementioned activities

  9. Cassette for handling banknotes or the like

    Science.gov (United States)

    Lundblad, Leif

    1981-08-11

    A cassette for banknotes and like valuable articles is provided with a displaceable lid (6) and locking means (10) for latching the lid of the cassette when the cassette is located outside a housing (25) in which it is intended to be placed. An operating means (8) is arranged to co-act with the locking means and with a latching element (15). The latching element is arranged to be released in dependence upon a pre-set program. A signal circuit is arranged to send a code signal to a detector circuit (23) when electrical contact elements on the cassette and the housing co-act with one another, which detector circuit, when the signal coincides with the signal program in the detector circuit, causes a signal to be sent for moving the latching means to a non-latching position.

  10. Current Understanding of Interactions between Nanoparticles and ABC Transporters in Cancer Cells.

    Science.gov (United States)

    Yin, Jian; Deng, Xudong; Zhang, Jie; Lin, Jun

    2018-03-14

    Adenosine triphosphate-binding cassette (ABC) transporters-mediated multidrug resistance (MDR) remains as a obstacle for effective cancer therapy. Nanoparticles (NPs)-based delivery systems are promising to overcome MDR, but only a few of them have been accepted for clinical treatment, due to characteristics such as insufficient transportation and potential toxicity. In this respect, mounting attention has been attracted towards interactions between NPs and ABC transporters, which hold a key role in the treatment of multidrug-resistant cancer and NP toxicity. In this review article, current knowledge on the involvement of ABC transporters in MDR and their inhibitors is provided. More importantly, recent literatures about the interactions between NPs and ABC transporters are summarized here. Organic and inorganic NPs inhibit the function of ABC transporters based on distinct mechanisms. The effects of organic NPs are caused by several excipients like surfactants, polymers, lipids and cyclodextrin, whereas inorganic NPs act as substrates of ABC transporters and competitively inhibit the efflux of drugs. Based on these interesting phenomena, a more thorough understanding of the specific mechanisms is necessary and essential in the hope to develop more efficient NPs to overcome MDR and decrease environmental toxicity of NPs. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  11. Curva ABC de fornecedores

    OpenAIRE

    Solano, Renato da Silva

    2003-01-01

    Dissertação (mestrado) - Universidade Federal de Santa Catarina, Centro Tecnológico. Programa de Pós-Graduação em Engenharia de Produção. Este trabalho faz uma revisão e busca dar uma contribuição aos gerentes de Planejamento, Programação, Controle e Gerenciamento de Empreendimentos e Obras através do uso das Curvas ABC. O trabalho utilizou 10 empreendimentos de classificação Alta (NBR 12721), incorporados por oito empresas do setor e fez uma análise dos projetos-padrão da NBR 12721. Foi f...

  12. Do You Know Your ABC?

    Science.gov (United States)

    Neale, Claire

    2013-01-01

    Within primary schools, the core subjects of literacy and numeracy are highly regarded, and rightly so, as children need to learn to read, write and be numerically literate. This means that all children learn their ABCs at an early age, But, what about the "other ABC"--"Airway, Breathing and Circulation?" Accidents and medical…

  13. ABC Technology Development Program

    International Nuclear Information System (INIS)

    1994-01-01

    The Accelerator-Based Conversion (ABC) facility will be designed to accomplish the following mission: 'Provide a weapon's grade plutonium disposition capability in a safe, economical, and environmentally sound manner on a prudent schedule for [50] tons of weapon's grade plutonium to be disposed on in [20] years.' This mission is supported by four major objectives: provide a reliable plutonium disposition capability within the next [15] years; provide a level of safety and of safety assurance that meets or exceeds that afforded to the public by modern commercial nuclear power plants; meet or exceed all applicable federal, state, and local regulations or standards for environmental compliance; manage the program in a cost effective manner. The ABC Technology Development Program defines the technology development activities that are required to accomplish this mission. The technology development tasks are related to the following topics: blanket system; vessel systems; reactivity control systems; heat transport system components; energy conversion systems; shutdown heat transport systems components; auxiliary systems; technology demonstrations - large scale experiments

  14. Human small cell lung cancer NYH cells selected for resistance to the bisdioxopiperazine topoisomerase II catalytic inhibitor ICRF-187 demonstrate a functional R162Q mutation in the Walker A consensus ATP binding domain of the alpha isoform

    DEFF Research Database (Denmark)

    Wessel, I; Jensen, L H; Jensen, P B

    1999-01-01

    -AMSA), which act by stabilizing enzyme-DNA-drug complexes at a stage in which the DNA gate strand is cleaved and the protein is covalently attached to DNA. Human small cell lung cancer NYH cells selected for resistance to ICRF-187 (NYH/187) showed a 25% increase in topoisomerase IIalpha level and no change......-clamp state in the enzyme's catalytic cycle caused by a decreased ATP binding by the mutated enzyme that is responsible for bisdioxopiperazine resistance....... demonstrated that R162Q conferred resistance to the bisdioxopiperazines ICRF-187 and -193 but not to etoposide or m-AMSA. Both etoposide and m-AMSA induced more DNA cleavage with purified R162Q enzyme than with the wt. The R162Q enzyme has a 20-25% decreased catalytic capacity compared to the wt and was almost...

  15. CryoEM and Molecular Dynamics of the Circadian KaiB-KaiC Complex Indicates That KaiB Monomers Interact with KaiC and Block ATP Binding Clefts

    Energy Technology Data Exchange (ETDEWEB)

    Villarreal, Seth A.; Pattanayek, Rekha; Williams, Dewight R.; Mori, Tetsuya; Qin, Ximing; Johnson, Carl H.; Egli, Martin; Stewart, Phoebe L. [Case Western; (Vanderbilt); (Vanderbilt-MED)

    2014-10-02

    The circadian control of cellular processes in cyanobacteria is regulated by a posttranslational oscillator formed by three Kai proteins. During the oscillator cycle, KaiA serves to promote autophosphorylation of KaiC while KaiB counteracts this effect. Here, we present a crystallographic structure of the wild-type Synechococcus elongatus KaiB and a cryo-electron microscopy (cryoEM) structure of a KaiBC complex. The crystal structure shows the expected dimer core structure and significant conformational variations of the KaiB C-terminal region, which is functionally important in maintaining rhythmicity. The KaiBC sample was formed with a C-terminally truncated form of KaiC, KaiC-Δ489, which is persistently phosphorylated. The KaiB–KaiC-Δ489 structure reveals that the KaiC hexamer can bind six monomers of KaiB, which form a continuous ring of density in the KaiBC complex. We performed cryoEM-guided molecular dynamics flexible fitting simulations with crystal structures of KaiB and KaiC to probe the KaiBC protein–protein interface. This analysis indicated a favorable binding mode for the KaiB monomer on the CII end of KaiC, involving two adjacent KaiC subunits and spanning an ATP binding cleft. A KaiC mutation, R468C, which has been shown to affect the affinity of KaiB for KaiC and lengthen the period in a bioluminescence rhythm assay, is found within the middle of the predicted KaiBC interface. The proposed KaiB binding mode blocks access to the ATP binding cleft in the CII ring of KaiC, which provides insight into how KaiB might influence the phosphorylation status of KaiC.

  16. Arabidopsis Transporter ABCG37/PDR9 contributes primarily highly oxygenated Coumarins to Root Exudation

    OpenAIRE

    Ziegler, J?rg; Schmidt, Stephan; Strehmel, Nadine; Scheel, Dierk; Abel, Steffen

    2017-01-01

    The chemical composition of root exudates strongly impacts the interactions of plants with microorganisms in the rhizosphere and the efficiency of nutrient acquisition. Exudation of metabolites is in part mediated by ATP-binding cassette (ABC) transporters. In order to assess the contribution of individual ABC transporters to root exudation, we performed an LC-MS based non-targeted metabolite profiling of semi-polar metabolites accumulating in root exudates of Arabidopsis thaliana plants and ...

  17. The chemistry of ABC

    Energy Technology Data Exchange (ETDEWEB)

    Jensen, R.J. [Los Alamos National Laboratory, NM (United States)

    1995-10-01

    ABC stand for accelerator based conversion of Pu. It is a unique approach to Pu destruction that allows for a well controlled and complete burn of Pu as may be required by treaty or policy. The central idea of the approach is to provide a spallation source of neutrons that allows the operation of a fissioning system without a critical mass and at a K effective less than one. Material to be fissioned is suspended in a molten salt medium for high temperature control and on-line removal of neutron absorbing fission products. This paper discusses the issues associated with the selection and operation of a molten salt chemical system: redox control, product removal, material feed, solubilities, deposition control, and a host of operational procedures.

  18. Uptake of L-cystine via an ABC transporter contributes defense of oxidative stress in the L-cystine export-dependent manner in Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Iwao Ohtsu

    Full Text Available Intracellular thiols like L-cystine and L-cystine play a critical role in the regulation of cellular processes. Here we show that Escherichia coli has two L-cystine transporters, the symporter YdjN and the ATP-binding cassette importer FliY-YecSC. These proteins import L-cystine, an oxidized product of L-cystine from the periplasm to the cytoplasm. The symporter YdjN, which is expected to be a new member of the L-cystine regulon, is a low affinity L-cystine transporter (Km = 1.1 μM that is mainly involved in L-cystine uptake from outside as a nutrient. E. coli has only two L-cystine importers because ΔydjNΔyecS mutant cells are not capable of growing in the minimal medium containing L-cystine as a sole sulfur source. Another protein YecSC is the FliY-dependent L-cystine transporter that functions cooperatively with the L-cystine transporter YdeD, which exports L-cystine as reducing equivalents from the cytoplasm to the periplasm, to prevent E. coli cells from oxidative stress. The exported L-cystine can reduce the periplasmic hydrogen peroxide to water, and then generated L-cystine is imported back into the cytoplasm via the ATP-binding cassette transporter YecSC with a high affinity to L-cystine (Km = 110 nM in a manner dependent on FliY, the periplasmic L-cystine-binding protein. The double disruption of ydeD and fliY increased cellular levels of lipid peroxides. From these findings, we propose that the hydrogen peroxide-inducible L-cystine/L-cystine shuttle system plays a role of detoxification of hydrogen peroxide before lipid peroxidation occurs, and then might specific prevent damage to membrane lipids.

  19. The ABCs of Sex Ed.

    Science.gov (United States)

    Sroka, Stephen R.

    2002-01-01

    Cites statistics on extent of sexually transmitted diseases and pregnancies among adolescents; describes ideological dispute over how to teach sex education; advocates teaching the ABCs of sex education: Abstinence, Be Monogamous, and Condoms. (PKP)

  20. Politseiniku lustlik ABC / Pekka Erelt

    Index Scriptorium Estoniae

    Erelt, Pekka, 1965-

    1999-01-01

    Politsei on välja andnud 'Politseiniku ABC', mis antakse igale politseinikule. Karikaturist Heiki Ernits on peaaegu igale taskuraamatu leheküljele joonistanud pildikese mundrimeestest kentsakates situatsioonides.

  1. ABC Transporters and Their Role in the Neoadjuvant Treatment of Esophageal Cancer

    Directory of Open Access Journals (Sweden)

    David Vrana

    2018-03-01

    Full Text Available The prognosis of esophageal cancer (EC is poor, despite considerable effort of both experimental scientists and clinicians. The tri-modality treatment consisting of neoadjuvant chemoradiation followed by surgery has remained the gold standard over decades, unfortunately, without significant progress in recent years. Suitable prognostic factors indicating which patients will benefit from this tri-modality treatment are missing. Some patients rapidly progress on the neoadjuvant chemoradiotherapy, which is thus useless and sometimes even harmful. At the same time, other patients achieve complete remission on neoadjuvant chemoradiotherapy and subsequent surgery may increase their risk of morbidity and mortality. The prognosis of patients ranges from excellent to extremely poor. Considering these differences, the role of drug metabolizing enzymes and transporters, among other factors, in the EC response to chemotherapy may be more important compared, for example, with pancreatic cancer where all patients progress on chemotherapy regardless of the treatment or disease stage. This review surveys published literature describing the potential role of ATP-binding cassette transporters, the genetic polymorphisms, epigenetic regulations, and phenotypic changes in the prognosis and therapy of EC. The review provides knowledge base for further research of potential predictive biomarkers that will allow the stratification of patients into defined groups for optimal therapeutic outcome.

  2. Maltose and maltodextrin utilization by Listeria monocytogenes depend on an inducible ABC transporter which is repressed by glucose.

    Directory of Open Access Journals (Sweden)

    Shubha Gopal

    2010-04-01

    Full Text Available In the environment as well as in the vertebrate intestine, Listeriae have access to complex carbohydrates like maltodextrins. Bacterial exploitation of such compounds requires specific uptake and utilization systems.We could show that Listeria monocytogenes and other Listeria species contain genes/gene products with high homology to the maltodextrin ABC transporter and utilization system of B. subtilis. Mutant construction and growth tests revealed that the L. monocytogenes gene cluster was required for the efficient utilization of maltodextrins as well as maltose. The gene for the ATP binding protein of the transporter was located distant from the cluster. Transcription analyses demonstrated that the system was induced by maltose/maltodextrins and repressed by glucose. Its induction was dependent on a LacI type transcriptional regulator. Repression by glucose was independent of the catabolite control protein CcpA, but was relieved in a mutant defective for Hpr kinase/phosphorylase.The data obtained show that in L. monocytogenes the uptake of maltodextrin and, in contrast to B. subtilis, also maltose is exclusively mediated by an ABC transporter. Furthermore, the results suggest that glucose repression of the uptake system possibly is by inducer exclusion, a mechanism not described so far in this organism.

  3. The ITER Divertor Cassette Project meeting

    International Nuclear Information System (INIS)

    Akiba, M.; Tivey, R.

    2000-01-01

    The Divertor Cassette Project topical meeting took place on April 5-7, 2000 at the JAERI Naka site in Japan. The meeting focused on the progress made by the three parties under task agreements on the development of carbon-fibre composite and tungsten armored high flux plasma-facing components

  4. ABC transporters in adaptive immunity.

    Science.gov (United States)

    Seyffer, Fabian; Tampé, Robert

    2015-03-01

    ABC transporters ubiquitously found in all kingdoms of life move a broad range of solutes across membranes. Crystal structures of four distinct types of ABC transport systems have been solved, shedding light on different conformational states within the transport process. Briefly, ATP-dependent flipping between inward- and outward-facing conformations allows directional transport of various solutes. The heterodimeric transporter associated with antigen processing TAP1/2 (ABCB2/3) is a crucial element of the adaptive immune system. The ABC transport complex shuttles proteasomal degradation products into the endoplasmic reticulum. These antigenic peptides are loaded onto major histocompatibility complex class I molecules and presented on the cell surface. We detail the functional modules of TAP, its ATPase and transport cycle, and its interaction with and modulation by other cellular components. In particular, we emphasize how viral factors inhibit TAP activity and thereby prevent detection of the infected host cell by cytotoxic T-cells. Merging functional details on TAP with structural insights from related ABC transporters refines the understanding of solute transport. Although human ABC transporters are extremely diverse, they still may employ conceptually related transport mechanisms. Appropriately, we delineate a working model of the transport cycle and how viral factors arrest TAP in distinct conformations. Deciphering the transport cycle of human ABC proteins is the major issue in the field. The defined peptidic substrate, various inhibitory viral factors, and its role in adaptive immunity provide unique tools for the investigation of TAP, making it an ideal model system for ABC transporters in general. This article is part of a Special Issue entitled Structural biochemistry and biophysics of membrane proteins. Copyright © 2014 Elsevier B.V. All rights reserved.

  5. Codon cassette mutagenesis: a general method to insert or replace individual codons by using universal mutagenic cassettes.

    OpenAIRE

    Kegler-Ebo, D M; Docktor, C M; DiMaio, D

    1994-01-01

    We describe codon cassette mutagenesis, a simple method of mutagenesis that uses universal mutagenic cassettes to deposit single codons at specific sites in double-stranded DNA. A target molecule is first constructed that contains a blunt, double-strand break at the site targeted for mutagenesis. A double-stranded mutagenic codon cassette is then inserted at the target site. Each mutagenic codon cassette contains a three base pair direct terminal repeat and two head-to-head recognition sequen...

  6. From MDR to MXR

    DEFF Research Database (Denmark)

    Litman, Thomas; Druley, T E; Stein, W D

    2001-01-01

    The ATP binding cassette (ABC) superfamily of membrane transporters is one of the largest protein classes known, and counts numerous proteins involved in the trafficking of biological molecules across cell membranes. The first known human ABC transporter was P-glycoprotein (P-gp), which confers......-gp, as well as other ABC transporters involved in MDR: the family of multidrug-resistance-associated proteins (MRP1-7), and the recently discovered ABC half-transporter MXR (also known as BCRP, ABCP and ABCG2). We describe recent progress in the analysis of protein structure-function relationships...

  7. Iowa ABC connections : [tech transfer summary].

    Science.gov (United States)

    2015-06-01

    The Iowa Department of Transportation (DOT) and other organizations have : been developing accelerated bridge construction (ABC) concepts, details, and : processes, and Iowa has come to be viewed as a national leader in the area of : ABC. However, th...

  8. Fuel cell cassette with compliant seal

    Science.gov (United States)

    Karl, Haltiner, Jr. J.; Anthony, Derose J.; Klotzbach, Darasack C.; Schneider, Jonathan R.

    2017-11-07

    A fuel cell cassette for forming a fuel cell stack along a fuel cell axis includes a cell retainer, a plate positioned axially to the cell retainer and defining a space axially with the cell retainer, and a fuel cell having an anode layer and a cathode layer separated by an electrolyte layer. The outer perimeter of the fuel cell is positioned in the space between the plate and the cell retainer, thereby retaining the fuel cell and defining a cavity between the cell retainer, the fuel cell, and the plate. The fuel cell cassette also includes a seal disposed within the cavity for sealing the edge of the fuel cell. The seal is compliant at operational temperatures of the fuel cell, thereby allowing lateral expansion and contraction of the fuel cell within the cavity while maintaining sealing at the edge of the fuel cell.

  9. The stability of cassette walls in compression

    Science.gov (United States)

    Voutay, Pierre-Arnaud

    Much research into the behaviour of cold formed steel columns in the last decade has focused on channel sections undergoing local, distortional and overall buckling. Light gauge steel cassette sections are a particular form of channel section which offers an alternative form of load-bearing wall assembly for use in low-rise steel framed construction. Cassette wall sections possess wide and slender flanges so that, by including intermediate stiffeners in these wide flanges, a significant increase in the ultimate load capacity may be achieved. However, the introduction of intermediate stiffeners also increases the number of buckling modes (stiffener buckling) and, therefore complicates the behaviour and increases the risk of interactive buckling between these modes. The work undertaken in this thesis aims to clarify the behaviour of wide flanges in compression with and without intermediate stiffeners. In this research, the distortional mode of web and narrow flange buckling was inhibited by connecting the narrow flanges of the cassettes together at suitable intervals. "Generalised Beam Theory" (GBT), which allows the individual buckling modes to be considered individually and in predetermined combinations, provides a particularly good tool with which to analyse and understand the buckling behaviour of cassette sections with and without intermediate stiffeners. "Generalised Beam Theory" (GBT) is used throughout this work to determine the elastic buckling stress of the sections studied (simply supported stiffened plates, as well as cassette sections). Since the economic design of cold-formed steel sections requires the consideration of post- buckling behaviour, elastic buckling values are not directly comparable with design code values which are usually based on the concept of effective width. Therefore, finite element analysis with both material and geometric nonlinearity has also been carried out in order to obtain the ultimate strength in the critical mode or mode

  10. Codon cassette mutagenesis: a general method to insert or replace individual codons by using universal mutagenic cassettes.

    Science.gov (United States)

    Kegler-Ebo, D M; Docktor, C M; DiMaio, D

    1994-05-11

    We describe codon cassette mutagenesis, a simple method of mutagenesis that uses universal mutagenic cassettes to deposit single codons at specific sites in double-stranded DNA. A target molecule is first constructed that contains a blunt, double-strand break at the site targeted for mutagenesis. A double-stranded mutagenic codon cassette is then inserted at the target site. Each mutagenic codon cassette contains a three base pair direct terminal repeat and two head-to-head recognition sequences for the restriction endonuclease Sapl, an enzyme that cleaves outside of its recognition sequence. The intermediate molecule containing the mutagenic cassette is then digested with Sapl, thereby removing most of the mutagenic cassette, leaving only a three base cohesive overhang that is ligated to generate the final insertion or substitution mutation. A general method for constructing blunt-end target molecules suitable for this approach is also described. Because the mutagenic cassette is excised during this procedure and alters the target only by introducing the desired mutation, the same cassette can be used to introduce a particular codon at all target sites. Each cassette can deposit two different codons, depending on the orientation in which it is inserted into the target molecule. Therefore, a series of eleven cassettes is sufficient to insert all possible amino acids at any constructed target site. Thus codon cassettes are 'off-the-shelf' reagents, and this methodology should be a particularly useful and inexpensive approach for subjecting multiple different positions in a protein sequence to saturation mutagenesis.

  11. The ABCs of particle physics

    CERN Document Server

    Biron, Lauren

    2016-01-01

    For lovers of rhymes and anthropomorphic Higgs bosons, Symmetry presents its first published board book, The ABCs of Particle Physics. Use it as an illustrated guide to basic particle- and astrophysics terms, or read it to your infant at bedtime, if you don’t mind their first word being “quark.”

  12. Therapeutic potential of inhibiting ABCE1 and eRF3 genes via siRNA strategy using chitosan nanoparticles in breast cancer cells

    Science.gov (United States)

    Cengiz, Bagdat Burcu; Asik, Mehmet Dogan; Kara, Goknur; Turk, Mustafa; Denkbas, Emir Baki

    2015-04-01

    In recent years, targeted cancer therapy strategies have begun to take the place of the conventional treatments. Inhibition of the specific genes, involved in cancer progress, via small interfering RNA (siRNA) has become one of the promising therapeutic approaches for cancer therapy. However, due to rapid nuclease degradation and poor cellular uptake of siRNA, a suitable carrier for siRNA penetration inside the cells is required. We used chitosan nanoparticles (CS-NPs) to efficiently deliver ATP-binding casette E1 (ABCE1) and eukaryotic release factor 3 (eRF3)-targeting siRNAs, individually and together, to reduce the proliferation and induce the apoptosis of breast cancer cells. The CS-NPs were generated by ionic gelation method using tripolyphosphate (TPP) as a crosslinker. Nanoparticles (NPs) were obtained with diameters ranging between 110 and 230 nm and the zeta potential of approximately 27 mV optimizing the solution pH to 4.5 and CS/TPP mass ratio to 3:1. Loading efficiencies of 98.69 % ± 0.051 and 98.83 % ± 0.047 were achieved when ABCE1 siRNA and eRF3 siRNA were entrapped into the NPs, respectively. Cell proliferation assay demonstrated that siRNA-loaded CS-NPs were more effective on cancer cells when compared to siRNAs without CS-NPs. Parallel results were also obtained by apoptosis/necrosis, double-staining analysis. Within our study, the potency of ABCE1 and eRF3 siRNAs were shown for the first time with this kind of polymeric delivery system. The results also indicated that ABCE1 and eRF3, important molecules in protein synthesis, could serve as effective targets to inhibit the cancer cells.

  13. Therapeutic potential of inhibiting ABCE1 and eRF3 genes via siRNA strategy using chitosan nanoparticles in breast cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Cengiz, Bagdat Burcu; Asik, Mehmet Dogan [Hacettepe University, Nanotechnology and Nanomedicine Division (Turkey); Kara, Goknur [Hacettepe University, Biochemistry Division, Chemistry Department (Turkey); Turk, Mustafa [Kirikkale University, Bioengineering Department (Turkey); Denkbas, Emir Baki, E-mail: denkbas@hacettepe.edu.tr [Hacettepe University, Biochemistry Division, Chemistry Department (Turkey)

    2015-04-15

    In recent years, targeted cancer therapy strategies have begun to take the place of the conventional treatments. Inhibition of the specific genes, involved in cancer progress, via small interfering RNA (siRNA) has become one of the promising therapeutic approaches for cancer therapy. However, due to rapid nuclease degradation and poor cellular uptake of siRNA, a suitable carrier for siRNA penetration inside the cells is required. We used chitosan nanoparticles (CS-NPs) to efficiently deliver ATP-binding casette E1 (ABCE1) and eukaryotic release factor 3 (eRF3)-targeting siRNAs, individually and together, to reduce the proliferation and induce the apoptosis of breast cancer cells. The CS-NPs were generated by ionic gelation method using tripolyphosphate (TPP) as a crosslinker. Nanoparticles (NPs) were obtained with diameters ranging between 110 and 230 nm and the zeta potential of approximately 27 mV optimizing the solution pH to 4.5 and CS/TPP mass ratio to 3:1. Loading efficiencies of 98.69 % ± 0.051 and 98.83 % ± 0.047 were achieved when ABCE1 siRNA and eRF3 siRNA were entrapped into the NPs, respectively. Cell proliferation assay demonstrated that siRNA-loaded CS-NPs were more effective on cancer cells when compared to siRNAs without CS-NPs. Parallel results were also obtained by apoptosis/necrosis, double-staining analysis. Within our study, the potency of ABCE1 and eRF3 siRNAs were shown for the first time with this kind of polymeric delivery system. The results also indicated that ABCE1 and eRF3, important molecules in protein synthesis, could serve as effective targets to inhibit the cancer cells.

  14. Disinfection Effect of Film Cassettes by Ultraviolet Irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Kweon, Dae Cheol; Park, Peom [Ajou Univ., Suwon (Korea, Republic of)

    2001-12-15

    A bacteria infection on film cassette contact surface was examined at the diagnostic radiology department. Studies have demonstrated a bactericidal effect of ultraviolet irradiation, and to assess the contamination level on film cassette contact surface as a predictor of patient prevent from nosocomial infection. The study showed that the laboratory result was identified non-pathologic and pathologic bacterial in the five different cassette size of the contact surface. Film cassettes were exposed to ultraviolet light for 1, 2 and 3 minutes. Ultraviolet light disinfection practices suitable for bacteria. The study concludes that presence of a bacterial infection will prevent a using antiseptic technique on film cassette contact surface. In conclusion, ultraviolet irradiate on film cassette over the surface more than 2 minutes. Ultraviolet dose of 1565 {mu}W {center_dot} s/cm{sup 2}Win in 30 second relative to ultraviolet dose in time.

  15. Disinfection Effect of Film Cassettes by Ultraviolet Irradiation

    International Nuclear Information System (INIS)

    Kweon, Dae Cheol; Park, Peom

    2001-01-01

    A bacteria infection on film cassette contact surface was examined at the diagnostic radiology department. Studies have demonstrated a bactericidal effect of ultraviolet irradiation, and to assess the contamination level on film cassette contact surface as a predictor of patient prevent from nosocomial infection. The study showed that the laboratory result was identified non-pathologic and pathologic bacterial in the five different cassette size of the contact surface. Film cassettes were exposed to ultraviolet light for 1, 2 and 3 minutes. Ultraviolet light disinfection practices suitable for bacteria. The study concludes that presence of a bacterial infection will prevent a using antiseptic technique on film cassette contact surface. In conclusion, ultraviolet irradiate on film cassette over the surface more than 2 minutes. Ultraviolet dose of 1565 μW · s/cm 2 Win in 30 second relative to ultraviolet dose in time

  16. A study on contamination and disinfection of film cassette

    International Nuclear Information System (INIS)

    Kweon, Dae Cheol; Chung, Kyung Mo; Choi, Ji Won

    2000-01-01

    In July 2000, a bacteria infection on film cassette contact surface was examined at the diagnostic radiology department of the S. hospital. The objective of this study was to assess the contamination level on film cassette contact surface as a predictor of patient to prevent from nosocomial infection. The study showed that the laboratory result was identified non-pathologic bacterial in the four different cassette size of the contact surface. The study concludes that presence of a bacterial infection will prevent a using antiseptic technique on film cassette contact surface. Also the education of nosocomial infection for radiographer will be required

  17. A study on contamination and disinfection of film cassette

    Energy Technology Data Exchange (ETDEWEB)

    Kweon, Dae Cheol; Chung, Kyung Mo [Seoul National University Hospital, Seoul (Korea, Republic of); Choi, Ji Won [University of Sydney, Sydney (Australia)

    2000-04-15

    In July 2000, a bacteria infection on film cassette contact surface was examined at the diagnostic radiology department of the S. hospital. The objective of this study was to assess the contamination level on film cassette contact surface as a predictor of patient to prevent from nosocomial infection. The study showed that the laboratory result was identified non-pathologic bacterial in the four different cassette size of the contact surface. The study concludes that presence of a bacterial infection will prevent a using antiseptic technique on film cassette contact surface. Also the education of nosocomial infection for radiographer will be required.

  18. Mobile gene cassettes: a fundamental resource for bacterial evolution.

    Science.gov (United States)

    Michael, Carolyn A; Gillings, Michael R; Holmes, Andrew J; Hughes, Lesley; Andrew, Nigel R; Holley, Marita P; Stokes, H W

    2004-07-01

    Horizontal gene transfer increases genetic diversity in prokaryotes to a degree not allowed by the limitations of reproduction by binary fission. The integron/gene cassette system is one of the most recently characterized examples of a system that facilitates horizontal gene transfer. This system, discovered in the context of multidrug resistance, is recognized in a clinical context for its role in allowing pathogens to adapt to the widespread use of antibiotics. Recent studies suggest that gene cassettes are common and encode functions relevant to many adaptive traits. To estimate the diversity of mobile cassettes in a natural environment, a molecular technique was developed to provide representative distributions of cassette populations at points within a sampling area. Subsequently, statistical methods analogous to those used for calculating species diversity were employed to assess the diversity of gene cassettes within the sample area in addition to gaining an estimate of cassette pool size. Results indicated that the number of cassettes within a 5x10-m sample area was large and that the overall mobile cassette metagenome was likely to be orders of magnitude larger again. Accordingly, gene cassettes appear to be capable of mobilizing a significant genetic resource and consequently have a substantial impact on bacterial adaptability.

  19. A Cassette Based System for Hydrogen Storage and Delivery

    Energy Technology Data Exchange (ETDEWEB)

    Britton Wayne E.

    2006-11-29

    A hydrogen storage system is described and evaluated. This is based upon a cassette, that is a container for managing hydrogen storage materials. The container is designed to be safe, modular, adaptable to different chemistries, inexpensive, and transportable. A second module receives the cassette and provides the necessary infrastructure to deliver hydrogen from the cassette according to enduser requirements. The modular concept has a number of advantages over approaches that are all in one stand alone systems. The advantages of a cassette based system are discussed, along with results from model and laboratory testing.

  20. Wireless technology for ABC Assessment

    DEFF Research Database (Denmark)

    Kristensen, Margit; Kyng, Morten; Christensen, Erika Frischknecht

    be just beside a victim to assess the ABC status. Consequently - in situations with many victims - monitoring equipment is only used for the (few) most seriously injured. Our ongoing research builds on data collected through extensive field studies and is described in several published papers. We......  In medical emergency service (EMS) the ABC status (Airway-Breathing-Circulation) of victims is essential in the decision making process regarding prioritizing and treatment. Today's use of information technology does, however, not inform directly about "A" and "B". The parameters are evaluated...... indirectly through assessment of collected biosensor data, such as ECG, and directly by the rescuer regarding how the patient looks and sounds. Moreover, the monitoring equipment used today is connected with displays by wires and thus only utilizes one display at a time. This means that the rescuers must...

  1. An ABC transporter B family protein, ABCB19, is required for cytoplasmic streaming and gravitropism of the inflorescence stems.

    Science.gov (United States)

    Okamoto, Keishi; Ueda, Haruko; Shimada, Tomoo; Tamura, Kentaro; Koumoto, Yasuko; Tasaka, Masao; Morita, Miyo Terao; Hara-Nishimura, Ikuko

    2016-01-01

    A significant feature of plant cells is the extensive motility of organelles and the cytosol, which was originally defined as cytoplasmic streaming. We suggested previously that a three-way interaction between plant-specific motor proteins myosin XIs, actin filaments, and the endoplasmic reticulum (ER) was responsible for cytoplasmic streaming. (1) Currently, however, there are no reports of molecular components for cytoplasmic streaming other than the actin-myosin-cytoskeleton and ER-related proteins. In the present study, we found that elongated cells of inflorescence stems of Arabidopsis thaliana exhibit vigorous cytoplasmic streaming. Statistical analysis showed that the maximal velocity of plastid movements is 7.26 µm/s, which is much faster than the previously reported velocities of organelles. Surprisingly, the maximal velocity of streaming in the inflorescence stem cells was significantly reduced to 1.11 µm/s in an Arabidopsis mutant, abcb19-101, which lacks ATP BINDING CASSETTE SUBFAMILY B19 (ABCB19) that mediates the polar transport of the phytohormone auxin together with PIN-FORMED (PIN) proteins. Polar auxin transport establishes the auxin concentration gradient essential for plant development and tropisms. Deficiency of ABCB19 activity eventually caused enhanced gravitropic responses of the inflorescence stems and abnormally flexed inflorescence stems. These results suggest that ABCB19-mediated auxin transport plays a role not only in tropism regulation, but also in cytoplasmic streaming.

  2. Loss of AtPDR11, a plasma membrane-localized ABC transporter, confers paraquat tolerance in Arabidopsis thaliana.

    Science.gov (United States)

    Xi, Jing; Xu, Ping; Xiang, Cheng-Bin

    2012-03-01

    Paraquat is one of the most widely used herbicides in the world. However, no paraquat transporter has been isolated in plants. Here, we describe paraquat-tolerant mutant pqt24-1, isolated from an activation-tagging library on the basis of its tolerance to 2 μm paraquat in the seedling stage. Molecular analysis revealed that the T-DNA was inserted in the 13th exon of At1g66950, which encodes AtPDR11, a member of the ATP-binding cassette transporter superfamily. As a result, AtPDR11 was knocked out in the mutant. Loss-of-function mutations of AtPDR11 led to reduced paraquat accumulation in plant cells. In addition, the AtPDR11 protein was specifically localized in the plasmalemma, suggesting AtPDR11 as a potential transporter of paraquat. This conclusion was supported by kinetic analysis of paraquat import. Further studies showed that the transcript level of AtPDR11 could be strongly induced by paraquat and other abiotic stresses including H(2) O(2) , indicating possible up-regulation of AtPDR11 expression by oxidative stress signaling. Thus, our data suggest that paraquat is an opportunistic substrate of AtPDR11 and the enhanced paraquat tolerance of pqt24-1 is due to reduced uptake of paraquat into plant cells. © 2011 The Authors. The Plant Journal © 2011 Blackwell Publishing Ltd.

  3. An ABC transporter B family protein, ABCB19, is required for cytoplasmic streaming and gravitropism of the inflorescence stems

    Science.gov (United States)

    Okamoto, Keishi; Ueda, Haruko; Shimada, Tomoo; Tamura, Kentaro; Koumoto, Yasuko; Tasaka, Masao; Morita, Miyo Terao; Hara-Nishimura, Ikuko

    2016-01-01

    Abstract A significant feature of plant cells is the extensive motility of organelles and the cytosol, which was originally defined as cytoplasmic streaming. We suggested previously that a three-way interaction between plant-specific motor proteins myosin XIs, actin filaments, and the endoplasmic reticulum (ER) was responsible for cytoplasmic streaming.1 Currently, however, there are no reports of molecular components for cytoplasmic streaming other than the actin-myosin-cytoskeleton and ER-related proteins. In the present study, we found that elongated cells of inflorescence stems of Arabidopsis thaliana exhibit vigorous cytoplasmic streaming. Statistical analysis showed that the maximal velocity of plastid movements is 7.26 µm/s, which is much faster than the previously reported velocities of organelles. Surprisingly, the maximal velocity of streaming in the inflorescence stem cells was significantly reduced to 1.11 µm/s in an Arabidopsis mutant, abcb19-101, which lacks ATP BINDING CASSETTE SUBFAMILY B19 (ABCB19) that mediates the polar transport of the phytohormone auxin together with PIN-FORMED (PIN) proteins. Polar auxin transport establishes the auxin concentration gradient essential for plant development and tropisms. Deficiency of ABCB19 activity eventually caused enhanced gravitropic responses of the inflorescence stems and abnormally flexed inflorescence stems. These results suggest that ABCB19-mediated auxin transport plays a role not only in tropism regulation, but also in cytoplasmic streaming. PMID:26337543

  4. INVENTORY CLASSIFICATION WITH ABC ANALYSIS

    OpenAIRE

    Kıyak, Erkan; Timuş, Oğuz Han; Karayel, Mehmet

    2015-01-01

    For a substantial organization, managing the expansive inventory is a serious problem. One of the methods to solve this problem used widely is classifying the inventory according to some criteria and managing inventory according to this classisifation. In this study, ABC classification methods are researched and Ng's model, which is one of the most widely used, selected for further investigation. An illustrative example is presented to show the usability of the Ng's method.

  5. Enterprise Architecture Data Pada Hotel ABC

    OpenAIRE

    Soesatyo, Stephanie; Wibowo, Adi; Handojo, Andreas

    2015-01-01

    Hotel ABC is a company engaged in hospitality management, i.e. room rental, rental of meeting rooms and coffee shop. Hotel ABC has a branch that has the same specifications of the hotel. In business process, Hotel ABC has information systems to support existing business processes. However, existing information systems have not been integrated as a whole. Based on the condition stated above, analysis and design of enterprise architecture information system is created for the company. The proce...

  6. myo-Inositol and d-Ribose Ligand Discrimination in an ABC Periplasmic Binding Protein

    Science.gov (United States)

    Herrou, Julien

    2013-01-01

    The periplasmic binding protein (PBP) IbpA mediates the uptake of myo-inositol by the IatP-IatA ATP-binding cassette transmembrane transporter. We report a crystal structure of Caulobacter crescentus IbpA bound to myo-inositol at 1.45 Å resolution. This constitutes the first structure of a PBP bound to inositol. IbpA adopts a type I PBP fold consisting of two α-β lobes that surround a central hinge. A pocket positioned between the lobes contains the myo-inositol ligand, which binds with submicromolar affinity (0.76 ± 0.08 μM). IbpA is homologous to ribose-binding proteins and binds d-ribose with low affinity (50.8 ± 3.4 μM). On the basis of IbpA and ribose-binding protein structures, we have designed variants of IbpA with inverted binding specificity for myo-inositol and d-ribose. Five mutations in the ligand-binding pocket are sufficient to increase the affinity of IbpA for d-ribose by 10-fold while completely abolishing binding to myo-inositol. Replacement of ibpA with these mutant alleles unable to bind myo-inositol abolishes C. crescentus growth in medium containing myo-inositol as the sole carbon source. Neither deletion of ibpA nor replacement of ibpA with the high-affinity ribose binding allele affected C. crescentus growth on d-ribose as a carbon source, providing evidence that the IatP-IatA transporter is specific for myo-inositol. This study outlines the evolutionary relationship between ribose- and inositol-binding proteins and provides insight into the molecular basis upon which these two related, but functionally distinct, classes of periplasmic proteins specifically bind carbohydrate ligands. PMID:23504019

  7. Specificity of drug transport mediated by CaMDR1: a major facilitator ...

    Indian Academy of Sciences (India)

    Unknown

    among these two tested drugs. Due to severe toxicity of these drugs to insect cells, further characterization of. CaMdr1p as a drug transporter could not be done with this system. Therefore, as an alternative, CaMdr1p and. Cdr1p, which is an ABC protein (ATP binding cassette) also involved in azole resistance in C. albicans, ...

  8. Insights into the Enhanced in vivo Fitness of Neisseria gonorrhoeae Driven by a Fluoroquinolone Resistance-Conferring Mutant DNA Gyrase

    Science.gov (United States)

    2015-02-05

    93 xi CHAPTER 1: Introduction Neisseria gonorrhoeae The majority of the 19 species within the genus Neisseria are...families: the resistance- nodulation-cell division (RND) family, the ATP-binding cassette (ABC) family, the small multi- drug resistance family, the...urine from men are the specimens of choice for NAAT-based detection of N. gonorrhoeae (23). Although not cleared by the Food and Drug Administration

  9. Abcd2 Is a Strong Modifier of the Metabolic Impairments in Peritoneal Macrophages of Abcd1-Deficient Mice

    OpenAIRE

    Muneer, Zahid; Wiesinger, Christoph; Voigtländer, Till; Werner, Hauke B.; Berger, Johannes; Forss-Petter, Sonja

    2014-01-01

    The inherited peroxisomal disorder X-linked adrenoleukodystrophy (X-ALD), associated with neurodegeneration and inflammatory cerebral demyelination, is caused by mutations in the ABCD1 gene encoding the peroxisomal ATP-binding cassette (ABC) transporter ABCD1 (ALDP). ABCD1 transports CoA-esters of very long-chain fatty acids (VLCFA) into peroxisomes for degradation by β-oxidation; thus, ABCD1 deficiency results in VLCFA accumulation. The closest homologue, ABCD2 (ALDRP), when overexpressed, c...

  10. Role of phosphoproteins involved in chemoresistance of colorectal cancer stem cells and immuno phenotypic comparative analysis

    International Nuclear Information System (INIS)

    Stassi, G.; Canzonieri, V.

    2009-01-01

    Recent studies demonstrated that colon cancers contain a cellular subpopulation, with stem cell-like proprieties, able to initiate and sustain tumour growth. These cells, so-called Cancer Initiating Cells (CICs), express the transmembrane antigen CD133. CD133 positive cells show slow proliferation rate, high expression of ABC (ATP-binding cassette) transporters and anti-apoptotic factors making them resistant to conventional therapies

  11. Transcriptional Analysis of Prebiotic Uptake and Catabolism by Lactobacillus acidophilus NCFM

    DEFF Research Database (Denmark)

    Andersen, Joakim Mark; Barrangou, Rodolphe; Abou Hachem, Maher

    2012-01-01

    and catabolism of 11 potential prebiotic compounds consisting of α- and β- linked galactosides and glucosides. These oligosaccharides induced genes encoding phosphoenolpyruvate-dependent sugar phosphotransferase systems (PTS), galactoside pentose hexuronide (GPH) permease, and ATP-binding cassette (ABC......-phospho-β-glucosidases implicated in the catabolism of gentiobiose and cellobiose. These findings highlight the broad oligosaccharide metabolic repertoire of L. acidophilus NCFM and establish a platform for selection and screening of both probiotic bacteria and prebiotic compounds that may positively...

  12. X-ray film cassette and method of making

    International Nuclear Information System (INIS)

    1980-01-01

    An x-ray film cassette which is capable of providing forces on the film that vary across the surface of the cassette is described. Methods of manufacture are discussed. The system is of particular use when large area films are used in conjunction with intensifying screens. (U.K.)

  13. Associations between dru Types and SCCmec Cassettes

    DEFF Research Database (Denmark)

    Bartels, Mette D; Boye, Kit; Oliveira, Duarte C

    2013-01-01

    types (dt) in 283 isolates, while eighteen isolates contained no dru repeats and one isolate resisted sequencing. The most common dru type, dt10a, was present in 53% of the sequenced isolates and was found in all SCCmec types, except type II. Seven (10%) of the 68 epidemiologically related patients had...... isolates with dru type variants indicating that dru typing is not useful as a first line epidemiological typing tool. However, MRSA isolates cultured from a single patient over a three year period exhibited a single dru type. The finding of dt10a in most SCCmec types suggests that dru and mecA originate......Molecular typing is an important tool in the investigation of methicillin resistant Staphylococcus aureus (MRSA) outbreaks and in following the evolution of MRSA. The staphylococcal cassette chromosome mec (SCCmec) contains a hypervariable region with a variable number of 40 bp repeats named direct...

  14. Playware ABC: Engineering Play for Everybody

    DEFF Research Database (Denmark)

    Lund, Henrik Hautop

    2017-01-01

    This paper describes the Playware ABC concept, and how it allows anybody, anywhere, anytime to be building bodies and brains, which facilitates users to construct, combine and create. The Playware ABC concept focuses engineering and IT system development on creating solutions that are usable by a...

  15. The ABC transporters in Candidatus Liberibacter asiaticus

    Science.gov (United States)

    Li, Wenlin; Cong, Qian; Pei, Jimin; Kinch, Lisa N; Grishin, Nick V

    2012-01-01

    Candidatus Liberibacter asiaticus (Ca. L. asiaticus) is a Gram-negative bacterium and the pathogen of Citrus Greening disease (Huanglongbing, HLB). As a parasitic bacterium, Ca. L. asiaticus harbors ABC transporters that play important roles in exchanging chemical compounds between Ca. L. asiaticus and its host. Here, we analyzed all the ABC transporter-related proteins in Ca. L. asiaticus. We identified 14 ABC transporter systems and predicted their structures and substrate specificities. In-depth sequence and structure analysis including multiple sequence alignment, phylogenetic tree reconstruction, and structure comparison further support their function predictions. Our study shows that this bacterium could use these ABC transporters to import metabolites (amino acids and phosphates) and enzyme cofactors (choline, thiamine, iron, manganese, and zinc), resist to organic solvent, heavy metal, and lipid-like drugs, maintain the composition of the outer membrane (OM), and secrete virulence factors. Although the features of most ABC systems could be deduced from the abundant experimental data on their orthologs, we reported several novel observations within ABC system proteins. Moreover, we identified seven nontransport ABC systems that are likely involved in virulence gene expression regulation, transposon excision regulation, and DNA repair. Our analysis reveals several candidates for further studies to understand and control the disease, including the type I virulence factor secretion system and its substrate that are likely related to Ca. L. asiaticus pathogenicity and the ABC transporter systems responsible for bacterial OM biosynthesis that are good drug targets. PMID:22807026

  16. ABC classification : service levels and inventory costs

    NARCIS (Netherlands)

    Teunter, R.H.; Babai, M.; Syntetos, A.

    2010-01-01

    ABC inventory classifications are widely used in practice, with demand value and demand volume as the most common ranking criteria. The standard approach in ABC applications is to set the same service level for all stock keeping units (SKUs) in a class. In this paper, we show (for three large real

  17. MicroRNA-299-3p promotes the sensibility of lung cancer to doxorubicin through directly targeting ABCE1.

    Science.gov (United States)

    Zheng, Dawei; Dai, Yan; Wang, Song; Xing, Xiaoyu

    2015-01-01

    MicroRNAs (miRNAs) are a class of endogenous, small non-coding RNAs which play important roles in various biological and cellular processes, including chemoresistance. The expression level of miR-299-3p was dysregulated in doxorubicin-resistance lung cancer cell lines. However, the exact role of miR-299-3p in doxorubicin-resistance is still unknown. In the present study, miR-299-3p was down-expressed in doxorubicin-resistant or -sensitive lung cancer samples and it was identified to directly targeted adenosine triphosphate binding cassette E1 (ABCE1) 3'-untranslated region (UTR) in lung cancer H69 cells by luciferase assay. After transfection of miR-299-3p mimics or ABCE1-siRNA, MTT assay confirmed that the H69/ADR cell proliferation was inhibited, as well as the enhanced cell inhibitory rate in the presence of doxorubicin. H69/ADR cell apoptosis rate was promoted after miR-299-3p or ABCE1-siRNA transfection. The results indicated that miR-299-3p promotes the sensibility of lung cancer to doxorubicin through suppression of ABCE1, at least partly. Therefore, the disordered decreased of miR-299-3p and resulting ABCE1 up-expression may contribute to chemoresistance of lung cancer, and miR-299-3p-ABCE1 may represent a new potential therapeutic target for the treatment of chemoresistance of lung cancer.

  18. Phylogenetic analysis of fungal ABC transporters

    Directory of Open Access Journals (Sweden)

    Driessen Arnold JM

    2010-03-01

    Full Text Available Abstract Background The superfamily of ABC proteins is among the largest known in nature. Its members are mainly, but not exclusively, involved in the transport of a broad range of substrates across biological membranes. Many contribute to multidrug resistance in microbial pathogens and cancer cells. The diversity of ABC proteins in fungi is comparable with those in multicellular animals, but so far fungal ABC proteins have barely been studied. Results We performed a phylogenetic analysis of the ABC proteins extracted from the genomes of 27 fungal species from 18 orders representing 5 fungal phyla thereby covering the most important groups. Our analysis demonstrated that some of the subfamilies of ABC proteins remained highly conserved in fungi, while others have undergone a remarkable group-specific diversification. Members of the various fungal phyla also differed significantly in the number of ABC proteins found in their genomes, which is especially reduced in the yeast S. cerevisiae and S. pombe. Conclusions Data obtained during our analysis should contribute to a better understanding of the diversity of the fungal ABC proteins and provide important clues about their possible biological functions.

  19. An ABC for decision making

    Energy Technology Data Exchange (ETDEWEB)

    Garcia, Luiz Henrique Costa, E-mail: luiz_mogi@yahoo.com.br [Associacao de Medicina Intensiva Brasileira (AMIB), Sao Paulo, SP (Brazil); Irmandade da Santa Casa de Misericordia de Sao Paulo, SP (Brazil); Ferreira, Bruna Cortez [Hospital de Base de Sao Jose do Rio Preto, SP (Brazil)

    2015-03-15

    The present study was aimed at proposing a systematic evaluation of cranial computed tomography, identifying the main aspects to be analyzed in order to facilitate the decision making process regarding diagnosis and management in emergency settings. The present descriptive study comprised a literature review at the following databases: Access Medicine and Access Emergency Medicine (McGraw-Hill Education); British Medical Journal Evidence Center; UptoDate; Bireme; PubMed; Lilacs; SciELO; ProQuest; Micromedex (Thomson Reuters); Embase. Once the literature review was completed, the authors identified the main diseases with tomographic repercussions and proposed the present system to evaluate cranial computed tomography images. An easy-to-memorize ABC system will facilitate the decision making in emergency settings, as it covers the main diseases encountered by intensivists and emergency physicians, and provides a sequential guidance about anatomical structures to be investigated as well as their respective alterations. (author)

  20. An ABC for decision making

    International Nuclear Information System (INIS)

    Garcia, Luiz Henrique Costa; Ferreira, Bruna Cortez

    2015-01-01

    The present study was aimed at proposing a systematic evaluation of cranial computed tomography, identifying the main aspects to be analyzed in order to facilitate the decision making process regarding diagnosis and management in emergency settings. The present descriptive study comprised a literature review at the following databases: Access Medicine and Access Emergency Medicine (McGraw-Hill Education); British Medical Journal Evidence Center; UptoDate; Bireme; PubMed; Lilacs; SciELO; ProQuest; Micromedex (Thomson Reuters); Embase. Once the literature review was completed, the authors identified the main diseases with tomographic repercussions and proposed the present system to evaluate cranial computed tomography images. An easy-to-memorize ABC system will facilitate the decision making in emergency settings, as it covers the main diseases encountered by intensivists and emergency physicians, and provides a sequential guidance about anatomical structures to be investigated as well as their respective alterations. (author)

  1. An ABC for decision making

    Directory of Open Access Journals (Sweden)

    Luiz Henrique Costa Garcia

    2015-04-01

    Full Text Available The present study was aimed at proposing a systematic evaluation of cranial computed tomography, identifying the main aspects to be analyzed in order to facilitate the decision making process regarding diagnosis and management in emergency settings. The present descriptive study comprised a literature review at the following databases: Access Medicine and Access Emergency Medicine (McGraw- Hill Education; British Medical Journal Evidence Center; UptoDate; Bireme; PubMed; Lilacs; SciELO; ProQuest; Micromedex (Thomson Reuters; Embase. Once the literature review was completed, the authors identified the main diseases with tomographic repercussions and proposed the present system to evaluate cranial computed tomography images. An easy-to-memorize ABC system will facilitate the decision making in emergency settings, as it covers the main diseases encountered by intensivists and emergency physicians, and provides a sequential guidance about anatomical structures to be investigated as well as their respective alterations.

  2. Observing cassette culture: user interface implications for digital music libraries

    OpenAIRE

    Toal, Jason

    2007-01-01

    Many people keep their collections of music on cassette tape even if they rarely listen to them. Images of these collections can be found online on photo sharing websites. What can we learn from such collections and what might they tell us about designing interfaces for new digital music libraries? The author conducts an online ethnographic study of over two hundred cassette tape collections, and over sixty participants with the aim of guiding future design of music collections. The author pr...

  3. Transcription, Processing, and Function of CRISPR Cassettes in Escherichia coli

    OpenAIRE

    Pougach, Ksenia; Semenova, Ekaterina; Bogdanova, Ekaterina; Datsenko, Kirill A.; Djordjevic, Marko; Wanner, Barry L.; Severinov, Konstantin

    2010-01-01

    CRISPR/Cas, bacterial and archaeal systems of interference with foreign genetic elements such as viruses or plasmids, consist of DNA loci called CRISPR cassettes (a set of variable spacers regularly separated by palindromic repeats) and associated cas genes. When a CRISPR spacer sequence exactly matches a sequence in a viral genome, the cell can become resistant to the virus. The CRISPR/Cas systems function through small RNAs originating from longer CRISPR cassette transcripts. While laborato...

  4. ABCs of Oral Health: Nutrition - Children

    Science.gov (United States)

    ... higher risk for heart attacks More ABCs of Oral Health A | B | C | D | E | F | G | H | ... Children Home | InfoBites | Find a Dentist | Your Family's Oral Health | RSS About AGD | Contact AGD | Site Map | Reprints ...

  5. ABC Inventory classification application: Özdemirler

    OpenAIRE

    Duru, Mehmet

    1989-01-01

    Ankara : The Department of Management and the Institute of Management Sciences of Bilkent Univ. , 1989. Thesis (Master's) -- Bilkent University, 1989. Includes bibliographical references leaves 37. ABC inventory classification can result in more effective control of business. In this work, ABC method is applied to Ozdemirler to examine the inventory profile of the store, and to be able to aid the management in allocating control effort among items more effectively. Dur...

  6. The yeast ABC transporter Pdr18 (ORF YNR070w) controls plasma membrane sterol composition, playing a role in multidrug resistance

    Science.gov (United States)

    Cabrito, Tânia R.; Teixeira, Miguel C.; Singh, Ashutosh; Prasad, Rajendra; Sá-Correia, Isabel

    2011-01-01

    The action of multidrug efflux pumps in MDR (multidrug resistance) acquisition has been proposed to partially depend on the transport of physiological substrates which may indirectly affect drug partition and transport across cell membranes. In the present study, the PDR18 gene [ORF (open reading frame) YNR070w], encoding a putative PDR (pleiotropic drug resistance) transporter of the ATP-binding cassette superfamily, was found to mediate plasma membrane sterol incorporation in yeast. The physiological role of Pdr18 is demonstrated to affect plasma membrane potential and is proposed to underlie its action as a MDR determinant, conferring resistance to the herbicide 2,4-D (2,4-dichlorophenoxyacetic acid). The action of Pdr18 in yeast tolerance to 2,4-D, which was found to contribute to reduce [14C]2,4-D intracellular accumulation, may be indirect, given the observation that 2,4-D exposure deeply affects the sterol plasma membrane composition, this effect being much stronger in a Δpdr18 background. PDR18 activation under 2,4-D stress is regulated by the transcription factors Nrg1, controlling carbon source availability and the stress response, and, less significantly, Yap1, involved in oxidative stress and MDR, and Pdr3, a key regulator of the yeast PDR network, consistent with a broad role in stress defence. Taken together, the results of the present study suggest that Pdr18 plays a role in plasma membrane sterol incorporation, this physiological trait contributing to an MDR phenotype. PMID:21831043

  7. An ABC transporter controls export of a Drosophila germ cell attractant.

    Science.gov (United States)

    Ricardo, Sara; Lehmann, Ruth

    2009-02-13

    Directed cell migration, which is critical for embryonic development, leukocyte trafficking, and cell metastasis, depends on chemoattraction. 3-hydroxy-3-methylglutaryl coenzyme A reductase regulates the production of an attractant for Drosophila germ cells that may itself be geranylated. Chemoattractants are commonly secreted through a classical, signal peptide-dependent pathway, but a geranyl-modified attractant would require an alternative pathway. In budding yeast, pheromones produced by a-cells are farnesylated and secreted in a signal peptide-independent manner, requiring the adenosine triphosphate-binding cassette (ABC) transporter Ste6p. Here we show that Drosophila germ cell migration uses a similar pathway, demonstrating that invertebrate germ cells, like yeast cells, are attracted to lipid-modified peptides. Components of this unconventional export pathway are highly conserved, suggesting that this pathway may control the production of similarly modified chemoattractants in organisms ranging from yeast to humans.

  8. AcEST: DK963275 [AcEST

    Lifescience Database Archive (English)

    Full Text Available trepto... 31 5.3 sp|P37276|DYHC_DROME Dynein heavy chain, cytoplasmic OS=Drosophi... 31 5.3 sp|P48410|ABCD1_...MOUSE ATP-binding cassette sub-family D member 1... 31 5.3 sp|P33897|ABCD1_HUMAN ...3 + KI E+ A+ET+ V+ ++ DWE KP GK Sbjct: 1230 QAKIVAEDKAVETRTVDFLN--DWEKTKPTGGK 1260 >sp|P48410|ABCD1...t: 663 LWKYHTHLLQFDGEGGWKFEKLDSAARLSLTEE 695 >sp|P33897|ABCD1_HUMAN ATP-binding cassette sub-family D member 1 OS=Homo sapiens GN=ABC...D1 PE=1 SV=2 Length = 745 Score = 30.8 bits (68), Expect = 5.3 Identities = 13/33 (

  9. Conformational cycle of the vitamin B12 ABC importer in liposomes detected by double electron-electron resonance (DEER).

    Science.gov (United States)

    Joseph, Benesh; Korkhov, Vladimir M; Yulikov, Maxim; Jeschke, Gunnar; Bordignon, Enrica

    2014-02-07

    Double electron-electron resonance is used here to investigate intermediates of the transport cycle of the Escherichia coli vitamin B12 ATP-binding cassette importer BtuCD-F. Previously, we showed the ATP-induced opening of the cytoplasmic gate I in TM5 helices, later confirmed by the AMP-PNP-bound BtuCD-F crystal structure. Here, other key residues are analyzed in TM10 helices (positions 307 and 322) and in the cytoplasmic gate II, i.e. the loop between TM2 and TM3 (positions 82 and 85). Without BtuF, binding of ATP induces detectable changes at positions 307 and 85 in BtuCD in liposomes. Together with BtuF, ATP triggers the closure of the cytoplasmic gate II in liposomes (reported by both positions 82 and 85). This forms a sealed cavity in the translocation channel in agreement with the AMP-PNP·BtuCD-F x-ray structure. When vitamin B12 and AMP-PNP are simultaneously present, the extent of complex formation is reduced, but the short 82-82 interspin distance detected indicates that the substrate does not affect the closed conformation of this gate. The existence of the BtuCD-F complex under these conditions is verified with spectroscopically orthogonal nitroxide and Gd(III)-based labels. The cytoplasmic gate II remains closed also in the vanadate-trapped state, but it reopens in the ADP-bound state of the complex. Therefore, we suggest that the substrate likely trapped in ATP·BtuCD-F can be released after ATP hydrolysis but before the occluded ADP-bound conformation is reached.

  10. Calibration of charcoal cassettes for radio-Iodine sampling

    International Nuclear Information System (INIS)

    Levinson, S.; Pelled, O.; Ballon, I.; Oved, S.; German, U.

    2004-01-01

    131 I is considered a high hazard radioisotope due to its abundance as a fission product, and its concentration in the thyroid gland. Monitoring 131 I in laboratories and determining its concentration in air is of great importance for Radiation Protection purposes. In order to achieve good collection efficiencies, monitoring devices are based on active charcoal cassettes, usually impregnated with TEDA 5% to enhance Iodine trapping (retention) efficiency. We employ at NRCN at the radio-iodine production laboratory continuous monitoring by air sampling through a cassette containing ∼26 gram activated coal, with a diameter of 57.4 mm and a height of 22 mm (TE2C 30x50 Mesh, manufactured by F and J., USA). A monitoring device, the RIS system, was described in the past (1). The charcoal cassette is replaced periodically, and the activity of the radio-Iodine is determined by gamma counting or spectrometry

  11. AcEST: BP913939 [AcEST

    Lifescience Database Archive (English)

    Full Text Available YMU001_000038_B01 468 Adiantum capillus-veneris mRNA. clone: YMU001_000038_B01. BP913939 - Show BP913939...is mRNA. clone: YMU001_000038_B01. Accession BP913939 Tissue type prothallium Developmental stage - Contig I...Nucleic Acids Res. 25:3389-3402. Query= BP913939|Adiantum capillus-veneris mRNA, ... F member 3... 86 9e-17 sp|Q66H39|ABCF3_RAT ATP-binding cassette sub-family F member 3 O... 85 1e-16 sp|Q5R9...aracterized ABC transporter ATP-binding... 56 6e-08 sp|P63390|YHES_ECO57 Uncharacterized ABC transporter ATP

  12. Blanket maintenance by remote means using the cassette blanket approach

    International Nuclear Information System (INIS)

    Werner, R.W.

    1978-01-01

    Induced radioactivity in the blanket and other parts of a fusion reactor close to the plasma zone will dictate remote assembly, disassembly, and maintenance procedures. Time will be of the essence in these procedures. They must be practicable and certain. This paper discusses the reduction of a complicated Tokamak reactor to a simpler assembly via the use of a vacuum building in which to house the reactor and the introduction in this new model of cassette blanket modules. The cassettes significantly simplify remote handling

  13. abc: An extensible AspectJ compiler

    DEFF Research Database (Denmark)

    Avgustinov, Pavel; Christensen, Aske Simon; Hendren, Laurie

    2005-01-01

    Research in the design of aspect-oriented programming languages requires a workbench that facilitates easy experimentation with new language features and implementation techniques. In particular, new features for AspectJ have been proposed that require extensions in many dimensions: syntax, type...... checking and code generation, as well as data flow and control flow analyses. The AspectBench Compiler (abc) is an implementation of such a workbench. The base version of abc implements the full AspectJ language. Its frontend is built, using the Polyglot framework, as a modular extension of the Java...... language. The use of Polyglot gives flexibility of syntax and type checking. The backend is built using the Soot framework, to give modular code generation and analyses. In this paper, we outline the design of abc, focusing mostly on how the design supports extensibility. We then provide a general overview...

  14. ABC model and the management of costs

    Directory of Open Access Journals (Sweden)

    Pravdić Predrag

    2016-01-01

    Full Text Available When a company has multiple objectives at the same time, they all must be considered and balanced when making any business decisions. Linking the markets, capital and resources so as to thus ensure the highest yield is, In fact, the search for competitive advantage as a basic condition for survival in a market economy. In highly detailed systems based on the management of costs or ABC (activity based costing systems, the cost of activities often result in erroneous evaluation of aggregate costs of the action. Improvements in information technology and monitoring decrease of technology costs enabled the ABC system to become a feasible system calculating costs in many organizations.

  15. ABC transporters : Lessons from a bacterial oligopeptide uptake system

    NARCIS (Netherlands)

    Doeven, Mark Koop

    2006-01-01

    ABC transporters are proteins that are involved in the uptake of nutrients and excretion of harmful substances in biological cells. Defects in human ABC transporters can result in severe diseases. Too high ABC transporter activity can lead to non-responsiveness of cancercells to chemotherapy, and

  16. Overcoming Multidrug Resistance in Cancer Stem Cells

    Directory of Open Access Journals (Sweden)

    Karobi Moitra

    2015-01-01

    Full Text Available The principle mechanism of protection of stem cells is through the expression of ATP-binding cassette (ABC transporters. These transporters serve as the guardians of the stem cell population in the body. Unfortunately these very same ABC efflux pumps afford protection to cancer stem cells in tumors, shielding them from the adverse effects of chemotherapy. A number of strategies to circumvent the function of these transporters in cancer stem cells are currently under investigation. These strategies include the development of competitive and allosteric modulators, nanoparticle mediated delivery of inhibitors, targeted transcriptional regulation of ABC transporters, miRNA mediated inhibition, and targeting of signaling pathways that modulate ABC transporters. The role of ABC transporters in cancer stem cells will be explored in this paper and strategies aimed at overcoming drug resistance caused by these particular transporters will also be discussed.

  17. ORF Alignment: NC_003212 [GENIUS II[Archive

    Lifescience Database Archive (English)

    Full Text Available ATP-binding protein [Desulfovibrio vulgaris subsp. ... vulgaris str. Hildenborough] gb|AAS96105.1| AB...C ... transporter, ATP-binding protein [Desulfovibrio vulgaris ... subsp. vulgaris str. Hilden... ... LQGEDLRKRFGQREVVRGVSVSVQQGEIVGLLGPNGAGKTTTFYMLTGIIK--PTAGIVR 58 ... Query: 124 LDAKMDILDMDEEMAERYLNEGFSGGEKKRNEILQLLMIEPKLAILDE

  18. ORF Alignment: NC_003210 [GENIUS II[Archive

    Lifescience Database Archive (English)

    Full Text Available ATP-binding protein [Desulfovibrio vulgaris subsp. ... vulgaris str. Hildenborough] gb|AAS96105.1| AB...C ... transporter, ATP-binding protein [Desulfovibrio vulgaris ... subsp. vulgaris str. Hilden... ... LQGEDLRKRFGQREVVRGVSVSVQQGEIVGLLGPNGAGKTTTFYMLTGIIK--PTAGIVR 58 ... Query: 124 LDAKMDILDMDEEMAERYLNEGFSGGEKKRNEILQLLMIEPKLAILDE

  19. ORF Alignment: NC_002973 [GENIUS II[Archive

    Lifescience Database Archive (English)

    Full Text Available ATP-binding protein [Desulfovibrio vulgaris subsp. ... vulgaris str. Hildenborough] gb|AAS96105.1| AB...C ... transporter, ATP-binding protein [Desulfovibrio vulgaris ... subsp. vulgaris str. Hilden... ... LQGEDLRKRFGQREVVRGVSVSVQQGEIVGLLGPNGAGKTTTFYMLTGIIK--PTAGIVR 58 ... Query: 124 LDAKMDILDMDEEMAERYLNEGFSGGEKKRNEILQLLMIEPKLAILDE

  20. The ABC-paradox: is Time Driven ABC relevant for small and Medium sized enterprises (SME)?

    DEFF Research Database (Denmark)

    Fladkjær, Henrik Find; Jensen, Erling

    -books and in curriculum at most business schools. But ABC is not used very much by the compa-nies. Especially in small and medium enterprises ABC is rarely implemented. This is primarily due to two things: First, the ABC model is too difficult to implement. Second, the data foundation for the model is not available...... in the required quality in the ERP systems used by the businesses. In this article we will explore ways of overcoming these two related problems. We will explore these issues particularly in relation to small and medium enterprises. Through a theoretical analysis and practical implementation in a medium sized...... manufacturing company we will analyse whether it is possible to create a satisfactory decision making foundation using a TDABC as a less complex cost model in the company. We have analysed our way to the conclusion that compared to ABC, the TDABC model provides the management with a number of pragmatic...

  1. Patterns of Availability and Use of Audiotape Cassettes in Special Libraries. Ph.D. Thesis

    Science.gov (United States)

    Hughes, J. M., II

    1975-01-01

    The availability and use of audiotape cassettes is studied in terms of user requirements. The following factors were examined: how special libraries utilize audiotape cassettes; who the users of the medium are; how the libraries acquire and maintain their collection; and opinions of librarians as to the value of the audiotape cassette as a medium for dissemination of information.

  2. Step 2: Know Your Diabetes ABCs

    Science.gov (United States)

    ... page please turn JavaScript on. Feature: Type 2 Diabetes Step 2: Know Your Diabetes ABCs Past Issues / Fall 2014 Table of Contents ... cholesterol helps remove cholesterol from your blood vessels. Diabetes HealthSense Find tools and programs that can help ...

  3. ABC – Advocates for Breast Cancer

    African Journals Online (AJOL)

    Cancer 2008;113(S8):221-224. [http://dx.doi.org/10.1002/cncr.23844]. ARTICLE. ABC – Advocates for Breast Cancer. A Steyn,1 Dip DA (Lond), LRAM (Speech and Drama) (Lond); J du Plessis;2 S Meyer,3 MSocSc (SW). 1 Immediate Past President, Reach to Recovery International, Queensland, Australia. 2 Chief Executive ...

  4. Phylogenetic analysis of fungal ABC transporters

    NARCIS (Netherlands)

    Kovalchuk, Andriy; Driessen, Arnold J. M.

    2010-01-01

    Background: The superfamily of ABC proteins is among the largest known in nature. Its members are mainly, but not exclusively, involved in the transport of a broad range of substrates across biological membranes. Many contribute to multidrug resistance in microbial pathogens and cancer cells. The

  5. Loss of plastoglobule kinases ABC1K1 and ABC1K3 causes conditional degreening, modified prenyl-lipids, and recruitment of the jasmonic acid pathway

    Science.gov (United States)

    Plastoglobules (PGs) are plastid lipid-protein particles. This study examines the function of PG-localized kinases ABC1K1 and ABC1K3 in Arabidopsis thaliana. Several lines of evidence suggested that ABC1K1 and ABC1K3 form a protein complex. Null mutants for both genes (abc1k1 and abc1k3) and the dou...

  6. Polycistronic transcription of fused cassettes and identification of translation initiation signals in an unusual gene cassette array from Pseudomonas aeruginosa [version 3; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Érica L. Fonseca

    2015-11-01

    Full Text Available The gene cassettes found in class 1 integrons are generally promoterless units composed by an open reading frame (ORF, a short 5’ untranslated region (UTR and a 3’ recombination site (attC. Fused gene cassettes are generated by partial or total loss of the attC from the first cassette in an array, creating, in some cases, a fusion with the ORF from the next cassette. These structures are rare and little is known about their mechanisms of mobilization and expression. The aim of this study was to evaluate the dynamic of mobilization and transcription of the gcu14-blaGES-1/aacA4 gene cassette array, which harbours a fused gene cassette represented by blaGES-1/aacA4. The cassette array was analyzed by Northern blot and real-time reverse transcription-polymerase chain reaction (RT-PCR in order to assess the transcription mechanism of blaGES-1/aacA4 fused cassette. Also, inverse polymerase chain reactions (PCR were performed to detect the free circular forms of gcu14, blaGES-1 and aacA4. The Northern blot and real time RT-PCR revealed a polycistronic transcription, in which the fused cassette blaGES-1/aacA4 is transcribed as a unique gene, while gcu14 (with a canonical attC recombination site has a monocistronic transcription. The gcu14 cassette, closer to the weak configuration of cassette promoter (PcW, had a higher transcription level than blaGES-1/aacA4, indicating that the cassette position affects the transcript amounts. The presence of ORF-11 at attI1, immediately preceding gcu14, and of a Shine-Dalgarno sequence upstream blaGES-1/aacA4 composes a scenario for the occurrence of array translation. Inverse PCR generated amplicons corresponding to gcu14, gcu14-aacA4 and gcu14-blaGES-1/aacA4 free circular forms, but not to blaGES-1 and aacA4 alone, indicating that the GES-1 truncated attC is not substrate of integrase activity and that these genes are mobilized together as a unique cassette. This study was original in showing the transcription

  7. Pouring and running a protein gel by reusing commercial cassettes.

    Science.gov (United States)

    Hwang, Alexander C; Grey, Paris H; Cuddy, Katrina; Oppenheimer, David G

    2012-02-12

    The evaluation of proteins using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis is a common technique used by biochemistry and molecular biology researchers. For laboratories that perform daily analyses of proteins, the cost of commercially available polyacrylamide gels (~$10/gel) can be considerable over time. To mitigate this cost, some researchers prepare their own polyacrylamide gels. Traditional methods of pouring these gels typically utilize specialized equipment and glass gel plates that can be expensive and preclude pouring many gels and storing them for future use. Furthermore, handling of glass plates during cleaning or gel pouring can result in accidental breakage creating a safety hazard, which may preclude their use in undergraduate laboratory classes. Our protocol demonstrates how to pour multiple protein gels simultaneously by recycling Invitrogen Nupage Novex minigel cassettes, and inexpensive materials purchased at a home improvement store. This economical and streamlined method includes a way to store the gels at 4°C for a few weeks. By re-using the plastic gel cassettes from commercially available gels, labs that run frequent protein gels can save significant costs and help the environment. In addition, plastic gel cassettes are extremely resistant to breakage, which makes them ideal for undergraduate laboratory classrooms.

  8. Plasmid-Encoded RepA Proteins Specifically Autorepress Individual repABC Operons in the Multipartite Rhizobium leguminosarum bv. trifolii Genome.

    Directory of Open Access Journals (Sweden)

    Kamil Żebracki

    Full Text Available Rhizobia commonly have very complex genomes with a chromosome and several large plasmids that possess genes belonging to the repABC family. RepA and RepB are members of the ParA and ParB families of partitioning proteins, respectively, whereas RepC is crucial for plasmid replication. In the repABC replicons, partitioning and replication functions are transcriptionally linked resulting in complex regulation of rep gene expression. The genome of R. leguminosarum bv. trifolii TA1 (RtTA1 consists of a chromosome and four plasmids (pRleTA1a-d, equipped with functional repABC genes. In this work, the regulation of transcription of the individual repABC cassettes of the four RtTA1 plasmids was studied. The involvement of the RepA and RepB as well as parS-like centromere sites in this process was depicted, demonstrating some dissimilarity in expression of respective rep regions. RtTA1 repABC genes of individual plasmids formed operons, which were negatively regulated by RepA and RepB. Individual RepA were able to bind to DNA without added nucleotides, but in the presence of ADP, bound specifically to their own operator sequences containing imperfect palindromes, and caused operon autorepression, whereas the addition of ATP stimulated non-specific binding of RepA to DNA. The RepA proteins were able to dimerize/oligomerize: in general dimers formed independently of ATP or ADP, although ATP diminished the concentration of oligomers that were produced. By the comprehensive approach focusing on a set of plasmids instead of individual replicons, the work highlighted subtle differences between the organization and regulation of particular rep operons as well as the structures and specificity of RepA proteins, which contribute to the fine-tuned coexistence of several replicons with similar repABC cassettes in the complex bacterial genome.

  9. Resistance to Bacillus thuringiensis Mediated by an ABC Transporter Mutation Increases Susceptibility to Toxins from Other Bacteria in an Invasive Insect

    Science.gov (United States)

    Zhang, Dandan; Gong, Lingling; He, Fei; Soberón, Mario; Bravo, Alejandra; Tabashnik, Bruce E.; Wu, Kongming

    2016-01-01

    Evolution of pest resistance reduces the efficacy of insecticidal proteins from the gram-positive bacterium Bacillus thuringiensis (Bt) used widely in sprays and transgenic crops. Recent efforts to delay pest adaptation to Bt crops focus primarily on combinations of two or more Bt toxins that kill the same pest, but this approach is often compromised because resistance to one Bt toxin causes cross-resistance to others. Thus, integration of Bt toxins with alternative controls that do not exhibit such cross-resistance is urgently needed. The ideal scenario of negative cross-resistance, where selection for resistance to a Bt toxin increases susceptibility to alternative controls, has been elusive. Here we discovered that selection of the global crop pest, Helicoverpa armigera, for >1000-fold resistance to Bt toxin Cry1Ac increased susceptibility to abamectin and spineotram, insecticides derived from the soil bacteria Streptomyces avermitilis and Saccharopolyspora spinosa, respectively. Resistance to Cry1Ac did not affect susceptibility to the cyclodiene, organophospate, or pyrethroid insecticides tested. Whereas previous work demonstrated that the resistance to Cry1Ac in the strain analyzed here is conferred by a mutation disrupting an ATP-binding cassette protein named ABCC2, the new results show that increased susceptibility to abamectin is genetically linked with the same mutation. Moreover, RNAi silencing of HaABCC2 not only decreased susceptibility to Cry1Ac, it also increased susceptibility to abamectin. The mutation disrupting ABCC2 reduced removal of abamectin in live larvae and in transfected Hi5 cells. The results imply that negative cross-resistance occurs because the wild type ABCC2 protein plays a key role in conferring susceptibility to Cry1Ac and in decreasing susceptibility to abamectin. The negative cross-resistance between a Bt toxin and other bacterial insecticides reported here may facilitate more sustainable pest control. PMID:26872031

  10. The Role of the Actin Cytoskeleton and Lipid Rafts in the Localization and Function of the ABCC1 Transporter

    Directory of Open Access Journals (Sweden)

    Jan Willem Kok

    2014-01-01

    Full Text Available ATP-binding cassette (ABC transporters are known to be important factors in multidrug resistance of tumor cells. Lipid rafts have been implicated in their localization in the plasma membrane, where they function as drug efflux pumps. This specific localization in rafts may support the activity of ABC/Abc transporters. This raises questions regarding the nature and composition of the lipid rafts that harbor ABC/Abc transporters and the dependence of ABC/Abc transporters—concerning their localization and activity—on lipid raft constituents. Here we review our work of the past 10 years aimed at evaluating whether ABC/Abc transporters are dependent on a particular membrane environment for their function. What is the nature of this membrane environment and which of the lipid raft constituents are important for this dependency? It turns out that cortical actin is of major importance for stabilizing the localization and function of the ABC/Abc transporter, provided it is localized in an actin-dependent subtype of lipid rafts, as is the case for human ABCC1/multidrug resistance-related protein 1 (MRP1 and rodent Abcc1/Mrp1 but not human ABCB1/P-glycoprotein (PGP. On the other hand, sphingolipids do not appear to be modulators of ABCC1/MRP1 (or Abcc1/Mrp1, even though they are coregulated during drug resistance development.

  11. Plant ABC proteins--a unified nomenclature and updated inventory.

    Science.gov (United States)

    Verrier, Paul J; Bird, David; Burla, Bo; Dassa, Elie; Forestier, Cyrille; Geisler, Markus; Klein, Markus; Kolukisaoglu, Uner; Lee, Youngsook; Martinoia, Enrico; Murphy, Angus; Rea, Philip A; Samuels, Lacey; Schulz, Burkhard; Spalding, Edgar J; Yazaki, Kazufumi; Theodoulou, Frederica L

    2008-04-01

    The ABC superfamily comprises both membrane-bound transporters and soluble proteins involved in a broad range of processes, many of which are of considerable agricultural, biotechnological and medical potential. Completion of the Arabidopsis and rice genome sequences has revealed a particularly large and diverse complement of plant ABC proteins in comparison with other organisms. Forward and reverse genetics, together with heterologous expression, have uncovered many novel roles for plant ABC proteins, but this progress has been accompanied by a confusing proliferation of names for plant ABC genes and their products. A consolidated nomenclature will provide much-needed clarity and a framework for future research.

  12. Disinfection efficacy of an ultraviolet light on film cassettes for preventive of the nosocomial infection

    Energy Technology Data Exchange (ETDEWEB)

    Kweon, Dae Cheol [Seoul National Univ. Hospital, Seoul (Korea, Republic of); Jeon, Yong Woong; Cho, Am [Dongguk Univ., Seoul (Korea, Republic of)

    2001-06-01

    The bacterial infection on film cassette contact surface was examined at the diagnostic radiology department of the S. hospital. The objective of this study was to assess the contamination level on film cassette contact surface as a predictor of patient prevention from nosocomial infection and for improvement of the hospital environment. The laboratory result was identified non-pathologic bacterial in the five different cassette size of the contact surface. Film cassettes were exposed to ultraviolet light for 1, 2 and 3 minutes. Ultraviolet light disinfection is proven suitable for bacterial. The study concludes that presence of a bacterial infection will prevent a using antiseptic technique on film cassette contact surface. In addition education of nosocomial infection for radiographers will be required. In conclusion, ultraviolet is considered effective to irradiate bacterial. Additionally, two minutes are required to sterilize film cassettes.

  13. Disinfection efficacy of an ultraviolet light on film cassettes for preventive of the nosocomial infection

    International Nuclear Information System (INIS)

    Kweon, Dae Cheol; Jeon, Yong Woong; Cho, Am

    2001-01-01

    The bacterial infection on film cassette contact surface was examined at the diagnostic radiology department of the S. hospital. The objective of this study was to assess the contamination level on film cassette contact surface as a predictor of patient prevention from nosocomial infection and for improvement of the hospital environment. The laboratory result was identified non-pathologic bacterial in the five different cassette size of the contact surface. Film cassettes were exposed to ultraviolet light for 1, 2 and 3 minutes. Ultraviolet light disinfection is proven suitable for bacterial. The study concludes that presence of a bacterial infection will prevent a using antiseptic technique on film cassette contact surface. In addition education of nosocomial infection for radiographers will be required. In conclusion, ultraviolet is considered effective to irradiate bacterial. Additionally, two minutes are required to sterilize film cassettes

  14. The role of the atypical kinases ABC1K7 and ABC1K8 in abscisic acid responses

    Directory of Open Access Journals (Sweden)

    Anna eManara

    2016-03-01

    Full Text Available The ABC1K family of atypical kinases (activity of bc1 complex kinase is represented in bacteria, archaea and eukaryotes. In plants they regulate diverse physiological processes in the chloroplasts and mitochondria, but their precise functions are poorly defined. ABC1K7 and ABC1K8 are probably involved in oxidative stress responses, isoprenyl lipid synthesis and distribution of iron within chloroplasts. Because reactive oxygen species take part in abscisic acid (ABA-mediated processes, we investigated the functions of ABC1K7 and ABC1K8 during germination, stomatal movement and leaf senescence. Both genes were upregulated by ABA treatment and some ABA-responsive physiological processes were affected in abc1k7 and abc1k8 mutants. Germination was more severely affected by ABA, osmotic stress and salt stress in the single and double mutants; the stomatal aperture was smaller in the mutants under standard growth conditions and was not further reduced by exogenous ABA application; ABA-induced senescence symptoms were more severe in the leaves of the single and double mutants compared to wild type leaves. Taken together, our results suggest that ABC1K7 and ABC1K8 might be involved in the cross-talk between ABA and ROS signaling.

  15. The x-ray source application test cassette for radiation exposures at the OMEGA laser

    Energy Technology Data Exchange (ETDEWEB)

    Fournier, K. B.; Rekow, V.; Emig, J. [Lawrence Livermore National Laboratory, Livermore, California 94551 (United States); Fisher, J. H.; Newlander, C. D. [Fifth Gait Technologies, Inc., Huntsville, Alabama 35803 (United States); Horton, R. [Gray Research, Inc., Huntsville, Alabama 35806 (United States); Davis, J. [Defense Threat Reduction Agency, Fort Belvoir, Virginia 22060 (United States)

    2012-10-15

    We have designed a sample cassette that can be used to position up to six samples in the OMEGA laser chamber. The cassette accommodates round samples up to 38.1 mm (1.5{sup Double-Prime }) in diameter and square samples up to 27 mm on a side, any of which can be up to 12.7 mm thick. Smaller specimens are centered with spacers. The test cassette allows each sample to have a unique filter scheme, with multiple filter regions in front of each sample. This paper will present mechanical design considerations and operational aspects of the x-ray source application cassette.

  16. On wiping the interior walls of 37-mm closed-face cassettes: an OSHA perspective.

    Science.gov (United States)

    Hendricks, Warren; Stones, Fern; Lillquist, Dean

    2009-12-01

    As early as 1976, Occupational Safety and Health Administration (OSHA) methods for analyzing metal samples collected using 37-mm polystyrene closed-face cassettes specified that any loose dust be transferred from the cassette to the digestion vessel, that the cassette be rinsed, and that, if necessary, the cassette be wiped out to help ensure that all particles that enter the cassette are included along with the filter as part of the sample for analysis. OSHA analytical methods for metal analysis were recently revised to explicitly require cassette wiping for all metal samples. This change was based on policy that any material entering the collection device constitutes part of the sample and on OSHA Salt Lake Technical Center research showing that invisible residue on the cassette walls can significantly contribute to the total sample results reported. OSHA procedures are consistent with guidance given in the NIOSH Manual of Analytical Methods. This guidance concludes that internal deposits in sampling cassettes should be included in the analysis and that one way to accomplish this would be to wipe or wash the internal surfaces of the cassette and include the material along with the filter for analysis.

  17. Nonlinear resonances in the ABC-flow

    Science.gov (United States)

    Didov, A. A.; Uleysky, M. Yu.

    2018-01-01

    In this paper, we study resonances of the ABC-flow in the near integrable case ( C ≪1 ). This is an interesting example of a Hamiltonian system with 3/2 degrees of freedom in which simultaneous existence of two resonances of the same order is possible. Analytical conditions of the resonance existence are received. It is shown numerically that the largest n :1 (n = 1, 2, 3) resonances exist, and their energies are equal to theoretical energies in the near integrable case. We provide analytical and numerical evidences for existence of two branches of the two largest n :1 (n = 1, 2) resonances in the region of finite motion.

  18. Conserved allosteric hot spots in the transmembrane domains of cystic fibrosis transmembrane conductance regulator (CFTR) channels and multidrug resistance protein (MRP) pumps.

    Science.gov (United States)

    Wei, Shipeng; Roessler, Bryan C; Chauvet, Sylvain; Guo, Jingyu; Hartman, John L; Kirk, Kevin L

    2014-07-18

    ATP-binding cassette (ABC) transporters are an ancient family of transmembrane proteins that utilize ATPase activity to move substrates across cell membranes. The ABCC subfamily of the ABC transporters includes active drug exporters (the multidrug resistance proteins (MRPs)) and a unique ATP-gated ion channel (cystic fibrosis transmembrane conductance regulator (CFTR)). The CFTR channel shares gating principles with conventional ligand-gated ion channels, but the allosteric network that couples ATP binding at its nucleotide binding domains (NBDs) with conformational changes in its transmembrane helices (TMs) is poorly defined. It is also unclear whether the mechanisms that govern CFTR gating are conserved with the thermodynamically distinct MRPs. Here we report a new class of gain of function (GOF) mutation of a conserved proline at the base of the pore-lining TM6. Multiple substitutions of this proline promoted ATP-free CFTR activity and activation by the weak agonist, 5'-adenylyl-β,γ-imidodiphosphate (AMP-PNP). TM6 proline mutations exhibited additive GOF effects when combined with a previously reported GOF mutation located in an outer collar of TMs that surrounds the pore-lining TMs. Each TM substitution allosterically rescued the ATP sensitivity of CFTR gating when introduced into an NBD mutant with defective ATP binding. Both classes of GOF mutations also rescued defective drug export by a yeast MRP (Yor1p) with ATP binding defects in its NBDs. We conclude that the conserved TM6 proline helps set the energy barrier to both CFTR channel opening and MRP-mediated drug efflux and that CFTR channels and MRP pumps utilize similar allosteric mechanisms for coupling conformational changes in their translocation pathways to ATP binding at their NBDs. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  19. ABC's "Person of the Week": American Values in Television News.

    Science.gov (United States)

    Larson, Stephanie Greco; Bailey, Martha

    1998-01-01

    Contributes to scholarship on values and ideologies in the media by analyzing five years of ABC's "Person of the Week" segments to identify prominent types of people and values endorsed by mainstream news media. Finds that individuals most frequently selected for ABC's honor lived in the United States; worked in politics and…

  20. Measuring Academic Behavioural Confidence: The ABC Scale Revisited

    Science.gov (United States)

    Sander, Paul; Sanders, Lalage

    2009-01-01

    The Academic Behavioural Confidence (ABC) scale has been shown to be valid and can be useful to teachers in understanding their students, enabling the design of more effective teaching sessions with large cohorts. However, some of the between-group differences have been smaller than expected, leading to the hypothesis that the ABC scale many not…

  1. Dashboard Auditing of Activity-Based Costing (ABC

    Directory of Open Access Journals (Sweden)

    Sorinel Capusneanu

    2009-03-01

    Full Text Available This article aims to define the dashboard auditing according to the specifics of Activity-Based Costing method (ABC. It describes the main objectives of dashboard auditing, the criteria that a dashboard auditor should meet and the step-by-step stages of the entire dashboard auditing process according to the Activity-Based Costing method (ABC.

  2. Dashboard Auditing of Activity-Based Costing (ABC)

    OpenAIRE

    Sorinel Capusneanu

    2009-01-01

    This article aims to define the dashboard auditing according to the specifics of Activity-Based Costing method (ABC). It describes the main objectives of dashboard auditing, the criteria that a dashboard auditor should meet and the step-by-step stages of the entire dashboard auditing process according to the Activity-Based Costing method (ABC).

  3. Dashboard auditing of ABC (Activity-Based Costing). Theoretical approaches

    OpenAIRE

    Căpuşneanu, Sorinel/I

    2009-01-01

    This article aims to define the dashboard auditing according to the specifics of Activity-Based Costing method (ABC). It describes the main objectives of dashboard auditing, the criteria that a dashboard auditor should meet and the step-by-step stages of the entire dashboard auditing process of an enterprise from steel industry according to the Activity-Based Costing method (ABC).

  4. Sustained Delivery of Chondroitinase ABC from Hydrogel System

    Directory of Open Access Journals (Sweden)

    Filippo Rossi

    2012-03-01

    Full Text Available In the injured spinal cord, chondroitin sulfate proteoglycans (CSPGs are the principal responsible of axon growth inhibition and they contribute to regenerative failure, promoting glial scar formation. Chondroitinase ABC (chABC is known for being able to digest proteoglycans, thus degrading glial scar and favoring axonal regrowth. However, its classic administration is invasive, infection-prone and clinically problematic. An agarose-carbomer (AC1 hydrogel, already used in SCI repair strategies, was here investigated as a delivery system capable of an effective chABC administration: the material ability to include chABC within its pores and the possibility to be injected into the target tissue were firstly proved. Subsequently, release kinetic and the maintenance of enzymatic activity were positively assessed: AC1 hydrogel was thus confirmed to be a feasible tool for chABC delivery and a promising device for spinal cord injury topic repair strategies.

  5. A survey of the radiographic cassettes disinfection of university hospitals in seoul

    International Nuclear Information System (INIS)

    Kweon, Dae Cheol; Park, Peom; Kim, Moon Sun; Kim, Dong Sung

    2001-01-01

    The purpose of this study is to prevent nosocomial infection in patients through contact of radiographic cassettes. Data were collected from radiographers working in 29 university hospitals in Seoul in February and March 2001. Radiographic cassettes were disinfected daily in 5 hospitals, weekly in 4 hospitals, monthly in 5 hospitals, bimonthly in 1 hospital and once every three months in another hospital. 12 other hospitals do not practice regular disinfections of radiographic cassettes. Gauze soaked in disinfectant solution is used in 7 hospitals while 11 hospitals used cotton and cloth soaked in disinfectant solution to clean the radiographic cassettes. 26 hospitals used 99% alcohol based disinfectant solutions while 3 hospitals used 75% alcohol based disinfectant, 26 hospitals use of intercourse cassettes outpatients and in patients. In 26 hospitals, all patients shared the same set of radiographic cassettes used in the hospitals, or in 26 hospitals, separate sets of radiographic cassettes are used for outpatients and inpatients. Separate sets of cassettes are used for ICU and inpatients in 6 others hospitals. 23 hospitals used the same sets of radiographic cassettes for all their patients. radiographic cassettes are cleaned in wash area in the study room of the radiographic department in 17 hospitals. 12 other hospitals do not have designated cleaning areas for the cassettes. All radiographers practiced hands washing with soap. All 29 hospitals surveyed have infection control committee. However, only 9 out of the 29 hospitals surveyed provided Infection · disinfections control education to radiographers. Only 3 hospitals have radiographers sitting in the infection control committee. Infection management education is conducted in 63 hospitals annually, twice a year in 1 hospital and once every 3 months in 2 hospitals

  6. An investigation of infection control for x-ray cassettes in a diagnostic imaging department

    Energy Technology Data Exchange (ETDEWEB)

    Fox, Matthew [School of Allied Health Professions and Science, Faculty of Health, Wellbeing and Science, University Campus Suffolk, Rope Walk, Ipswich, Suffolk, IP4 1LT (United Kingdom); Harvey, Jane M. [School of Allied Health Professions and Science, Faculty of Health, Wellbeing and Science, University Campus Suffolk, Rope Walk, Ipswich, Suffolk, IP4 1LT (United Kingdom)], E-mail: j.harvey@ucs.ac.uk

    2008-11-15

    Introduction: This research was conducted to investigate if X-ray cassettes could be a possible source of pathogens capable of causing nosocomial infections, and if they could be a possible vector for cross infection within the hospital environment. Method: The research involved the swabbing of X-ray cassettes in a Diagnostic Imaging Department of a large hospital in the east of England. Two areas of the Diagnostic Imaging Department were included in the study. Research concentrated on X-ray cassettes used for mobile radiography, accident and emergency and inpatient use. Forty cassettes were swabbed in total specifically for general levels of bacterial contamination, also for the presence or absence of methicillin-resistant Staphylococcus aureus (MRSA). A mapping exercise was completed following the location of an X-ray cassette typically used in mobile radiography. The exercise noted the level of direct contact with patient's skin and other possible routes of infection. Results: The results demonstrated that there were large levels of growth of samples taken from cassettes and developed in the Microbiology Department. Coagulase-negative Staphylococcus, Micrococci, Diptheroids and species of Bacillus were all identified. The mapping exercise in which the journey of a 35/43 cm cassette used for mobile radiography was tracked found that contact with patient's skin and potential pathogens or routes of cross infection was a common occurrence whilst undertaking mobile radiography. Conclusion: The research has identified the presence of bacterial contamination on cassettes. The research established that X-ray cassettes/imaging plates are often exposed to pathogens and possible routes of cross infection; also that patient's skin often comes directly in contact with the X-ray cassette/imaging plate. The research also shows that as cassettes/imaging plates are a potential source of cross infection, the Diagnostic Imaging Department may be partly responsible

  7. An investigation of infection control for x-ray cassettes in a diagnostic imaging department

    International Nuclear Information System (INIS)

    Fox, Matthew; Harvey, Jane M.

    2008-01-01

    Introduction: This research was conducted to investigate if X-ray cassettes could be a possible source of pathogens capable of causing nosocomial infections, and if they could be a possible vector for cross infection within the hospital environment. Method: The research involved the swabbing of X-ray cassettes in a Diagnostic Imaging Department of a large hospital in the east of England. Two areas of the Diagnostic Imaging Department were included in the study. Research concentrated on X-ray cassettes used for mobile radiography, accident and emergency and inpatient use. Forty cassettes were swabbed in total specifically for general levels of bacterial contamination, also for the presence or absence of methicillin-resistant Staphylococcus aureus (MRSA). A mapping exercise was completed following the location of an X-ray cassette typically used in mobile radiography. The exercise noted the level of direct contact with patient's skin and other possible routes of infection. Results: The results demonstrated that there were large levels of growth of samples taken from cassettes and developed in the Microbiology Department. Coagulase-negative Staphylococcus, Micrococci, Diptheroids and species of Bacillus were all identified. The mapping exercise in which the journey of a 35/43 cm cassette used for mobile radiography was tracked found that contact with patient's skin and potential pathogens or routes of cross infection was a common occurrence whilst undertaking mobile radiography. Conclusion: The research has identified the presence of bacterial contamination on cassettes. The research established that X-ray cassettes/imaging plates are often exposed to pathogens and possible routes of cross infection; also that patient's skin often comes directly in contact with the X-ray cassette/imaging plate. The research also shows that as cassettes/imaging plates are a potential source of cross infection, the Diagnostic Imaging Department may be partly responsible for adding to

  8. The ABC of ABC : An analysis of attribute-based credentials in the light of data protection, privacy and identity.

    NARCIS (Netherlands)

    Korenhof, P.E.I.; Koning, Merel; Alpár, Gergely; Hoepman, J.H.; Padullés, Joan Balcells; i Martínez, Agustí Cerrillo; Poch, Miquel Peguera; López, Ismael Peña; de Moner, María José Pifarré; Solana, Mònica Vilasau

    2014-01-01

    Our networked society increasingly needs secure identity sys- tems. The Attribute-based credential (ABC) technology is designed to be privacy-friendlier than contemporary authentication methods, which often suffer from information leakage. So far, however, some of the wider implications of ABC have

  9. Transcriptional interactions suggest niche segregation among microorganisms in the human gut

    DEFF Research Database (Denmark)

    Plichta, Damian Rafal; Juncker, Agnieszka; dos Santos, Marcelo Bertalan Quintanilha

    2016-01-01

    functional and metabolic interactions between cohabiting species 2,3. To move beyond species co-occurrence networks, we systematically identified transcriptional interactions between pairs of coexisting gut microbes using metagenomics and microarray-based metatranscriptomics data from 233 stool samples from...... biosynthesis, ATP-binding cassette (ABC) transporters, flagella assembly and bacterial chemotaxis, as well as for the metabolism of carbohydrates, amino acids and cofactors. The analysis gives the first insight into the microbial community-wide transcriptional interactions, and suggests that the regulation...

  10. Insights on the evolution of mycoparasitism from the genome of Clonostachys rosea

    DEFF Research Database (Denmark)

    Karlsson, Magnus; Durling, Mikael Brandström; Choi, Jaeyoung

    2015-01-01

    Clonostachys rosea is a mycoparasitic fungus that can control several important plant diseases. Here we report on the genome sequencing of C. rosea and a comparative genome analysis, in order to resolve the phylogenetic placement of C. rosea and to study the evolution of mycoparasitism as a fungal...... of gene family evolution reveals several distinct differences between the included mycoparasites. C. rosea contains significantly more ATP-binding cassette (ABC) transporters, polyketide synthases, cytochrome P450 monooxygenases, pectin lyases, glucose-methanol-choline oxidoreductases and lytic...

  11. Combined Transcriptome and Proteome Analysis of Bifidobacterium animalis subsp. lactis BB-12 Grown on Xylo-Oligosaccharides and a Model of Their Utilization

    DEFF Research Database (Denmark)

    Gilad, Ofir; Jacobsen, Susanne; Stuer-Lauridsen, B.

    2010-01-01

    of these (beta-D-xylosidase, sugar-binding protein, and xylose isomerase) showed higher abundance on XOS. Based on the obtained results, a model for the catabolism of XOS in BB-12 is suggested, according to which the strain utilizes an ABC (ATP-binding cassette) transport system (probably for oligosaccharides...... XOS or glucose. The analyses show that 9 of the 10 genes that encode proteins predicted to play a role in XOS catabolism (i.e., XOS-degrading and -metabolizing enzymes, transport proteins, and a regulatory protein) were induced by XOS at the transcriptional level, and the proteins encoded by three...

  12. Overcoming Multidrug Resistance in Human Cancer Cells by Natural Compounds

    Directory of Open Access Journals (Sweden)

    Tomohiro Nabekura

    2010-05-01

    Full Text Available Multidrug resistance is a phenomenon whereby tumors become resistant to structurally unrelated anticancer drugs. P-glycoprotein belongs to the large ATP-binding cassette (ABC transporter superfamily of membrane transport proteins. P-glycoprotein mediates resistance to various classes of anticancer drugs including vinblastine, daunorubicin, and paclitaxel, by actively extruding the drugs from the cells. The quest for inhibitors of anticancer drug efflux transporters has uncovered natural compounds, including (--epigallocatechin gallate, curcumin, capsaicin, and guggulsterone, as promising candidates. In this review, studies on the effects of natural compounds on P-glycoprotein and anticancer drug efflux transporters are summarized.

  13. Conserved ABC Transport System Regulated by the General Stress Response Pathways of Alpha- and Gammaproteobacteria

    Energy Technology Data Exchange (ETDEWEB)

    Herrou, Julien; Willett, Jonathan W.; Czyż, Daniel M.; Babnigg, Gyorgy; Kim, Youngchang; Crosson, Sean; Stock, Ann M.

    2016-12-19

    ABSTRACT

    Brucella abortusσE1is an EcfG family sigma factor that regulates the transcription of dozens of genes in response to diverse stress conditions and is required for maintenance of chronic infection in a mouse model. A putative ATP-binding cassette transporter operon,bab1_0223-bab1_0226, is among the most highly activated gene sets in the σE1regulon. The proteins encoded by the operon resemble quaternary ammonium-compatible solute importers but are most similar in sequence to the broadly conserved YehZYXW system, which remains largely uncharacterized. Transcription ofyehZYXWis activated by the general stress sigma factor σSinEnterobacteriaceae, which suggests a functional role for this transport system in bacterial stress response across the classesAlphaproteobacteriaandGammaproteobacteria. We present evidence thatB. abortusYehZYXW does not function as an importer of known compatible solutes under physiological conditions and does not contribute to the virulence defect of a σE1-null strain. The solein vitrophenotype associated with genetic disruption of this putative transport system is reduced growth in the presence of high Li+ion concentrations. A crystal structure ofB. abortusYehZ revealed a class II periplasmic binding protein fold with significant structural homology toArchaeoglobus fulgidusProX, which binds glycine betaine. However, the structure

  14. Cassette blanket and vacuum building: key elements in fusion reactor maintenance

    International Nuclear Information System (INIS)

    Werner, R.W.

    1977-01-01

    The integration of two concepts important to fusion power reactors is discussed. The first concept is the vacuum building which improves upon the current fusion reactor designs. The second concept, the use of the cassette blanket within the vacuum building environment, introduces four major improvements in blanket design: cassette blanket module, zoning concept, rectangular blanket concept, and internal tritium recovery

  15. Evolutionary Origin of the Staphylococcal Cassette Chromosome mec (SCCmec)

    DEFF Research Database (Denmark)

    Rolo, Joana; Worning, Peder; Nielsen, Jesper Boye

    2017-01-01

    Several lines of evidence indicate that the most primitive staphylococcal species, those of the Staphylococcus sciuri group, were involved in the first stages of evolution of the staphylococcal cassette chromosome mec (SCCmec), the genetic element carrying the β-lactam resistance gene mecA. However......, many steps are still missing from this evolutionary history. In particular, it is not known how mecA was incorporated into the mobile element SCC prior to dissemination among Staphylococcus aureus and other pathogenic staphylococcal species. To gain insights into the possible contribution of several...... species of the Staphylococcus sciuri group to the assembly of SCCmec, we sequenced the genomes of 106 isolates, comprising S. sciuri (n = 76), Staphylococcus vitulinus (n = 18), and Staphylococcus fleurettii (n = 12) from animal and human sources, and characterized the native location of mecA and the SCC...

  16. A two-cassette reporter system for assessing target gene translation and target gene product inclusion body formation

    DEFF Research Database (Denmark)

    2016-01-01

    The present invention relates to a dual cassette reporter system capable of assessing target gene translation and target gene product folding. The present invention further relates to vectors and host cells comprising the dual cassette reporter system. In addition the invention relates to the use...... of the dual cassette reporter system for assessing target gene translation and target gene product folding....

  17. Context-driven discovery of gene cassettes in mobile integrons using a computational grammar.

    Science.gov (United States)

    Tsafnat, Guy; Coiera, Enrico; Partridge, Sally R; Schaeffer, Jaron; Iredell, Jon R

    2009-09-08

    Gene discovery algorithms typically examine sequence data for low level patterns. A novel method to computationally discover higher order DNA structures is presented, using a context sensitive grammar. The algorithm was applied to the discovery of gene cassettes associated with integrons. The discovery and annotation of antibiotic resistance genes in such cassettes is essential for effective monitoring of antibiotic resistance patterns and formulation of public health antibiotic prescription policies. We discovered two new putative gene cassettes using the method, from 276 integron features and 978 GenBank sequences. The system achieved kappa = 0.972 annotation agreement with an expert gold standard of 300 sequences. In rediscovery experiments, we deleted 789,196 cassette instances over 2030 experiments and correctly relabelled 85.6% (alpha > or = 95%, E analysis demonstrated that for 72,338 missed deletions, two adjacent deleted cassettes were labeled as a single cassette, increasing performance to 94.8% (mean sensitivity = 0.92, specificity = 1, F-score = 0.96). Using grammars we were able to represent heuristic background knowledge about large and complex structures in DNA. Importantly, we were also able to use the context embedded in the model to discover new putative antibiotic resistance gene cassettes. The method is complementary to existing automatic annotation systems which operate at the sequence level.

  18. Context-driven discovery of gene cassettes in mobile integrons using a computational grammar

    Directory of Open Access Journals (Sweden)

    Schaeffer Jaron

    2009-09-01

    Full Text Available Abstract Background Gene discovery algorithms typically examine sequence data for low level patterns. A novel method to computationally discover higher order DNA structures is presented, using a context sensitive grammar. The algorithm was applied to the discovery of gene cassettes associated with integrons. The discovery and annotation of antibiotic resistance genes in such cassettes is essential for effective monitoring of antibiotic resistance patterns and formulation of public health antibiotic prescription policies. Results We discovered two new putative gene cassettes using the method, from 276 integron features and 978 GenBank sequences. The system achieved κ = 0.972 annotation agreement with an expert gold standard of 300 sequences. In rediscovery experiments, we deleted 789,196 cassette instances over 2030 experiments and correctly relabelled 85.6% (α ≥ 95%, E ≤ 1%, mean sensitivity = 0.86, specificity = 1, F-score = 0.93, with no false positives. Error analysis demonstrated that for 72,338 missed deletions, two adjacent deleted cassettes were labeled as a single cassette, increasing performance to 94.8% (mean sensitivity = 0.92, specificity = 1, F-score = 0.96. Conclusion Using grammars we were able to represent heuristic background knowledge about large and complex structures in DNA. Importantly, we were also able to use the context embedded in the model to discover new putative antibiotic resistance gene cassettes. The method is complementary to existing automatic annotation systems which operate at the sequence level.

  19. Downregulation of eIF4G by microRNA-503 enhances drug sensitivity of MCF-7/ADR cells through suppressing the expression of ABC transport proteins.

    Science.gov (United States)

    Pan, Xia; Yang, Xiaoyan; Zang, Jinglei; Zhang, Si; Huang, Nan; Guan, Xinxin; Zhang, Jianhua; Wang, Zhihui; Li, Xi; Lei, Xiaoyong

    2017-06-01

    Overexpression of adenosine triphosphate-binding cassette (ABC) transport protein is emerging as a critical contributor to anticancer drug resistance. The eukaryotic translation initiation factor (eIF) 4F complex, the key modulator of mRNA translation, is regulated by the phosphoinositide 3-kinase-AKT-mammalian target of rapamycin pathway in anticancer drug-resistant tumors. The present study demonstrated the roles of ABC translation protein alterations in the acquisition of the Adriamycin (ADM)-resistant phenotype of MCF-7 human breast cells. Quantitative polymerase chain reaction and western blot analysis were applied to examine the differences in mRNA and protein levels, respectively. It was found that the expression of the ABC sub-family B member 1, ABC sub-family C member 1 and ABC sub-family G member 2 transport proteins were upregulated in MCF-7/ADR cells. An MTT assay was used to detect the cell viability, from the results MCF-7/ADR cells were less sensitive to ADM, tamoxifen (TAM) and taxol (TAX) treatment compared with MCF-7 cells. We predicted that the 3'-untranslated region of eukaryotic translation initiation factor 4-γ 1 (eIF4G) contains a potential miRNA binding site for microRNA (miR)-503 through using computational programs. These binding sites were confirmed by luciferase reporter assays. eIF4G mRNA degradation was accelerated in cells transfected with miR-503 mimics. Furthermore, it was demonstrated that eIF4G and ABC translation proteins were significantly downregulated in MCF-7/ADR cells after transfection with miR-503. It was found that miR-503 mimics could sensitize the cells to treatment with ADM, TAM and TAX. These findings demonstrated for the first time that eIF4G acted as a key factor in MCF-7/ADR cells, and may be an efficient agent for preventing and reversing multi-drug resistance in breast cancer.

  20. Physiological function of ABCG1.

    Science.gov (United States)

    Ito, Toshimitsu

    2003-10-01

    Since ATP-binding cassette transporter A1 (ABCA1) was discovered as the cause of Tangier disease and familial high-density lipoprotein (HDL) deficiency, many investigators have been interested in the relationship between ABC transporters and the mechanism underlying abnormal lipid metabolism. ABCG1 is an ABC half transporter that facilitates efflux excess cholesterol from macrophages. To elucidate the potential physiological role of ABCG1, we have initiated a series of studies overexpressing ABCG1, using an adenovirus vector (rABCG1-Adv) in C57BL mice. Overexpression of ABCG1 in the liver of mice using recombinant ABCG1 vectors results in decreased plasma HDL levels and increased biliary cholesterol excretion, and indicates that ABCG1 can modulate plasma lipoprotein levels in vivo. ABCG1 and the other ABC transporters might play an important role in cholesterol homeostasis, especially in the liver.

  1. ABCG2/BCRP decreases the transfer of a food-born chemical carcinogen, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in perfused term human placenta

    International Nuclear Information System (INIS)

    Myllynen, Paeivi; Kummu, Maria; Kangas, Tiina; Ilves, Mika; Immonen, Elina; Rysae, Jaana; Pirilae, Rauna; Lastumaeki, Anni; Vaehaekangas, Kirsi H.

    2008-01-01

    We have studied the role of ATP binding cassette (ABC) transporters in fetal exposure to carcinogens using 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) a known substrate for ABC transporters as a model compound. In perfusion of human term placenta, transfer of 14 C-PhIP (2 μM) through the placenta resulted in fetal-to-maternal concentration ratio (FM ratio) of 0.72 ± 0.09 at 6 h. The specific ABCG2 inhibitor KO143 increased the transfer of 14 C-PhIP from maternal to fetal circulation (FM ratio 0.90 ± 0.08 at 6 h, p 14 C-PhIP (R = - 0.81, p 14 C-PhIP in perfused human placenta. Also, PhIP may modify ABC transporter expression in choriocarinoma cells

  2. Circumvention of multi-drug resistance of cancer cells by Chinese herbal medicines

    Directory of Open Access Journals (Sweden)

    Lin Ge

    2010-07-01

    Full Text Available Abstract Multi-drug resistance (MDR of cancer cells severely limits therapeutic outcomes. A proposed mechanism for MDR involves the efflux of anti-cancer drugs from cancer cells, primarily mediated by ATP-binding cassette (ABC membrane transporters including P-glycoprotein. This article reviews the recent progress of using active ingredients, extracts and formulae from Chinese medicine (CM in circumventing ABC transporters-mediated MDR. Among the ABC transporters, Pgp is the most extensively studied for its role in MDR reversal effects. While other MDR reversal mechanisms remain unclear, Pgp inhibition is a criterion for further mechanistic study. More mechanistic studies are needed to fully establish the pharmacological effects of potential MDR reversing agents.

  3. Circumvention of multi-drug resistance of cancer cells by Chinese herbal medicines.

    Science.gov (United States)

    Chai, Stella; To, Kenneth Kw; Lin, Ge

    2010-07-25

    Multi-drug resistance (MDR) of cancer cells severely limits therapeutic outcomes. A proposed mechanism for MDR involves the efflux of anti-cancer drugs from cancer cells, primarily mediated by ATP-binding cassette (ABC) membrane transporters including P-glycoprotein. This article reviews the recent progress of using active ingredients, extracts and formulae from Chinese medicine (CM) in circumventing ABC transporters-mediated MDR. Among the ABC transporters, Pgp is the most extensively studied for its role in MDR reversal effects. While other MDR reversal mechanisms remain unclear, Pgp inhibition is a criterion for further mechanistic study. More mechanistic studies are needed to fully establish the pharmacological effects of potential MDR reversing agents.

  4. ABC's of Being Smart: I Can "C" Clearly Now

    Science.gov (United States)

    Foster, Joanne

    2011-01-01

    In this paper, the author focuses on C of the ABC's of being smart. She continues to categorize the points for readers. These categories include the following: (1) being; (2) doing; and (3) stretching.

  5. Compilation of accelerated bridge construction (ABC) bridges : final report.

    Science.gov (United States)

    2016-01-01

    Development of accelerated bridge construction (ABC) technologies has been occurring across the country, many times in : isolation. Although FHWA and others have worked to facilitate communication between these efforts, there was not a : comprehensiv...

  6. ABC-B transporter genes in Dirofilaria immitis.

    Science.gov (United States)

    Bourguinat, Catherine; Che, Hua; Mani, Thangadurai; Keller, Kathy; Prichard, Roger K

    2016-08-01

    Dirofilaria immitis is a filarial nematode causing infection and heartworm disease in dogs and other canids, cats, and occasionally in humans. Prevention with macrocyclic lactones (ML) is recommended during the mosquito transmission season. Recently, ML resistance has been reported. ABC-B transporter genes are thought to be involved in the mechanism of ML resistance in other nematodes. This study aimed to identify all the ABC-B transporter genes in D. immitis using as a reference the nDi.2.2 D. immitis whole genome, which is not completely annotated. Using bioinformatic tools and PCR amplification on pooled D. immitis genomic DNA and on pooled cDNA, nine ABC transporter genes including one pseudogene were characterized. Bioinformatic and phylogenetic analyses allowed identification of three P-glycoproteins (Pgps) (Dim-pgp-3 Dim-pgp-10, Dim-pgp-11), of two ABC-B half transporter genes (one ortholog of Cel-haf-4 and Cel-haf-9; and one ortholog of Cel-haf-1 and Cel-haf-3), of one ABC half transporter gene (ortholog of Cel-haf-5) that contained an ABC-C motif, and of one additional half transporter that would require functional study for characterization. The number of ABC-B transporter genes identified was lower than in Caenorhabditis elegans and Haemonchus contortus. Further studies are needed to understand their possible role in ML resistance in D. immitis. These ABC transporters constitute a base for ML resistance investigation in D. immitis and advance our understanding of the molecular biology of this parasite. Copyright © 2016. Published by Elsevier Ltd.

  7. Application of the ABC analysis in management of food inventory

    OpenAIRE

    Radek Toušek

    2009-01-01

    The article is focused on managing stock inventory with help of the ABC method. The theory is explained on an actual company dealing with retail grocery distribution within the south bohemian region. The ABC method is applied on 4,405 inventory items regarding their 2008 year turnover. Results of the analysis are processed into a system plan to reduce the asset liability and decrease the stock inventory while maintaining the requested quality of logistic services.

  8. Radiation exposure reduction by use of Kevlar cassettes in the neonatal nursery.

    Science.gov (United States)

    Herman, M W; Mak, H K; Lachman, R S

    1987-05-01

    A study was performed to determine whether the use of Kevlar cassettes in the neonatal intensive care nursery would reduce radiation exposure to patients. The radiation dose to the neonates was measured by using thermoluminescent dosimeters. In addition, the attenuation of the Kevlar cassettes and the sensitivity of the film-screen combination were compared with the previously used system. The greatest radiation reduction using a mobile X-ray unit was 27%; based on sensitivity measurements, the theoretical reduction averaged 38%. The reduction in radiation exposure resulted from reduced attenuation by the Kevlar cassette.

  9. A clinical trial of a rare earth screen/film system in a periapical cassette

    Energy Technology Data Exchange (ETDEWEB)

    Kogon, S.L.; Stephens, R.G.; Reid, J.A.; Lubus, N.J.

    1984-04-01

    In a clinical trial, a slow rare earth screen/film system (Siemens Titan 2D/Kodak XG) was used to obtain intraoral radiographs at conventional monitoring stages in endodontic treatment. The screen film image proved to be an effective substitute for the direct-exposure Ultraspeed periapical film. The intraoral cassettes, designed and fabricated for the study, were an adaptation of the flexible, vacuum-sealed cassettes used in mammography. It is believed that when a practicable periapical cassette is manufactured, many additional indications for the system are probable. Major reductions in patient exposure of at least 85% to 90% per periapical film would be effected.

  10. A clinical trial of a rare earth screen/film system in a periapical cassette

    International Nuclear Information System (INIS)

    Kogon, S.L.; Stephens, R.G.; Reid, J.A.; Lubus, N.J.

    1984-01-01

    In a clinical trial, a slow rare earth screen/film system (Siemens Titan 2D/Kodak XG) was used to obtain intraoral radiographs at conventional monitoring stages in endodontic treatment. The screen film image proved to be an effective substitute for the direct-exposure Ultraspeed periapical film. The intraoral cassettes, designed and fabricated for the study, were an adaptation of the flexible, vacuum-sealed cassettes used in mammography. It is believed that when a practicable periapical cassette is manufactured, many additional indications for the system are probable. Major reductions in patient exposure of at least 85% to 90% per periapical film would be effected

  11. ABC transporter research: going strong 40 years on.

    Science.gov (United States)

    Theodoulou, Frederica L; Kerr, Ian D

    2015-10-01

    In most organisms, ABC transporters constitute one of the largest families of membrane proteins. In humans, their functions are diverse and underpin numerous key physiological processes, as well as being causative factors in a number of clinically relevant pathologies. Advances in our understanding of these diseases have come about through combinations of genetic and protein biochemical investigations of these transporters and the power of in vitro and in vivo investigations is helping to develop genotype-phenotype understanding. However, the importance of ABC transporter research goes far beyond human biology; microbial ABC transporters are of great interest in terms of understanding virulence and drug resistance and industrial biotechnology researchers are exploring the potential of prokaryotic ABC exporters to increase the capacity of synthetic biology systems. Plant ABC transporters play important roles in transport of hormones, xenobiotics, metals and secondary metabolites, pathogen responses and numerous aspects of development, all of which are important in the global food security area. For 3 days in Chester, this Biochemical Society Focused Meeting brought together researchers with diverse experimental approaches and with different fundamental questions, all of which are linked by the commonality of ABC transporters. © 2015 Authors.

  12. Use of an improved simultaneous tomography cassette in linear tomography

    International Nuclear Information System (INIS)

    Egender, G.; Pirker, E.; Gornik, E.; Innsbruck Univ.

    1984-01-01

    An improved simultaneous tomography cassette according to P. Landau was tried out for four months using four tomographs in routine work. The mode of operation is based on accurate control of the relative speeds of the individual x-ray films resulting in simultaneous imaging of 6 equidistant tomographic levels. Clinical testing was effected in 80 cases: nephrotomography, of the lungs, the hilum, and the skeleton. In particular, the article describes imaging of the renal arteries by simultaneous tomography for the purpose of finding out the cause of hypertension, and if there is suspicion of a space-occupying growth in the kidney, basing on the urogram. The specific advantages of this technique are, on the one hand, improved diagnostic efficiency (the tomograms are taken during the same respiratory phase, more rapid diagnosis especially with accident patients), and, on the other hand, an important reduction in the x-ray exposure of the patient; furthermore, the life of the x-ray tube is prolonged, and there is a definite saving of time for both patient and personnel, the image quality being comparable with that of single-layer tomography. (orig.) [de

  13. Resistance-Gene Cassettes Associated With Salmonella enterica Genotypes.

    Science.gov (United States)

    Bakhshi, Bita; Ghafari, Mohsen; Pourshafie, Mohammad R; Zarbakhsh, Behnaz; Katouli, Mohammad; Rahbar, Mohammad; Hajia, Masoud; Hosseini-Aliabad, Neda; Boustanshenas, Mina

    2015-01-01

    The epidemiology of salmonellosis is complex because of the diversity and different serotypes of Salmonella enterica (S. enterica) that occur in different reservoirs and geographic incidences. To determine the genotype distribution and resistance-gene content of 2 classes of integron among S. enterica isolates. Thirty-six S. enterica species were isolated and tested for their serological distribution and the resistance-gene contents of 2 classes of integron, as well as for their genetic diversity, using the pulsed-field gel electrophoresis (PFGE) genotyping method. Serogroups E (36.1%) and D (30.5%) were dominant among the isolates. All of the isolates in serogroup D belonged to the serovar enteritidis. The aadA1 gene was found within all resistance-gene cassettes. We observed 4 common and 26 single pulsotypes among the isolates, which indicated a high degree of genetic diversity among the isolates. Using the PulseNet International standard protocol, it was found that these isolates were different from those reported previously in Iran. The presence of a few common and new pulsotypes among the isolates suggests the emergence and spread of new clones of S. enterica in Iran. Copyright© by the American Society for Clinical Pathology (ASCP).

  14. Optimal ABC inventory classification using interval programming

    Science.gov (United States)

    Rezaei, Jafar; Salimi, Negin

    2015-08-01

    Inventory classification is one of the most important activities in inventory management, whereby inventories are classified into three or more classes. Several inventory classifications have been proposed in the literature, almost all of which have two main shortcomings in common. That is, the previous methods mainly rely on an expert opinion to derive the importance of the classification criteria which results in subjective classification, and they need precise item parameters before implementing the classification. While the problem has been predominantly considered as a multi-criteria, we examine the problem from a different perspective, proposing a novel optimisation model for ABC inventory classification in the form of an interval programming problem. The proposed interval programming model has two important features compared to the existing methods: it provides optimal results instead of an expert-based classification and it does not require precise values of item parameters, which are not almost always available before classification. Finally, by illustrating the proposed classification model in the form of numerical example, conclusion and suggestions for future works are presented.

  15. amdSYM, a new dominant recyclable marker cassette for Saccharomyces cerevisiae

    NARCIS (Netherlands)

    Solis-Escalante, D.; Kuijpers, N.G.A.; Bongaerts, N.; Bolat, I.; Bosman, L.; Pronk, J.T.; Daran, J.M.; Daran-Lapujade, P.A.S.

    2012-01-01

    Despite the large collection of selectable marker genes available for Saccharomyces cerevisiae, marker availability can still present a hurdle when dozens of genetic manipulations are required. Recyclable markers, counterselectable cassettes that can be removed from the targeted genome after use,

  16. Influence of cassette design on three-dimensional perfusion culture of artificial bone.

    Science.gov (United States)

    Du, Dajiang; Ushida, Takashi; Furukawa, Katsuko S

    2015-01-01

    Media perfusion is often required to maintain cell viability within topographically complex 3-dimensional scaffold cultures. Osteoblast-seeded scaffolds for bone regeneration require robust cell proliferation and survival both within the scaffold and over the exterior for optimal osteogenic capacity. Conventional press-fitting cassettes ensure internal fluid flow through the scaffold but may restrict external flow around the scaffold, resulting in a barren (cell-free) external scaffold surface. In this study, we aimed to solve this problem by modifying the cassette structure to enhance external flow in an oscillatory perfusion culture system. Mouse osteoblast-like MC 3T3-E1 cells were seeded in porous ceramic scaffolds and incubated for 3 days either under static culture conditions or in an oscillatory perfusion bioreactor. Scaffolds were held in the bioreactor with either conventional press-fitting cassettes or cassettes with rings to separate the scaffold exterior from the internal cassette wall. The external surfaces of scaffolds maintained under static conditions were well seeded, but cells failed to grow deeply into the core, reflecting poor internal chemotransport. Alternatively, scaffolds cultured by perfusion with press-fitting cassettes had poor cell viability at the cassette-external scaffold surface interface, but cells were widely distributed within the scaffold core. Scaffolds cultured using the modified cassettes with 1 or 2 rings exhibited uniformly distributed living cells throughout the internal pores and over the entire external surface, possibly because of the improved medium flow over the scaffold surface. This modified oscillatory perfusion culture system may facilitate the production of engineered bone with superior osteogenic capacity for grafting. © 2014 Wiley Periodicals, Inc.

  17. Design, simplified cloning, and in-silico analysis of multisite small interfering RNA-targeting cassettes

    OpenAIRE

    Baghban-Kohnehrouz, Bahram; Nayeri, Shahnoush

    2016-01-01

    Multiple gene silencing is being required to target and tangle metabolic pathways in eukaryotes and researchers have to develop a subtle method for construction of RNA interference (RNAi) cassettes. Although, several vectors have been developed due to different screening and cloning strategies but still some potential limitations remain to be dissolved. Here, we worked out a simple cloning strategy to develop multisite small interfering RNA (siRNA) cassette from different genes by two cloning...

  18. New Integron-Associated Gene Cassette Encoding a 3-N-Aminoglycoside Acetyltransferase

    OpenAIRE

    Levings, Renee S.; Partridge, Sally R.; Lightfoot, Diane; Hall, Ruth M.; Djordjevic, Steven P.

    2005-01-01

    A fifth gene cassette containing an aacC gene, aacCA5, was found in an aacCA5-aadA7 cassette array in a class 1 integron isolated from a multiply drug resistant Salmonella enterica serovar Kentucky strain. The AacC-A5 or AAC(3)-Ie acetyltransferase encoded by aacCA5 is related to other AAC(3)-I enzymes and confers resistance to gentamicin.

  19. Metodologia ABC: implantação numa microempresa ABC methodology: implementation on a micro firm

    Directory of Open Access Journals (Sweden)

    Orlando Duran

    2000-08-01

    Full Text Available Este trabalho relata a implantação da metodologia de custos baseados em atividades numa microempresa do ramo metalúrgico. A proposta pretende demonstrar a viabilidade de aplicar esta técnica em empresas sem importar seu tamanho, só realizando algumas adaptações que garantam baixo investimento e curto espaço de tempo para obter os resultados. Na parte final do trabalho se realiza uma análise dos resultados obtidos verificando-se o potencial da informação gerada pela metodologia e seu uso como ferramenta de gestão.This paper presents an implementation of the activity based costing (ABC methodology in a small firm. The approach presented is intended to demonstrate the feasibility of applying the ABC methodology at any sized firm, only through few adaptations for ensuring low investments fees and speed in obtaining results and information from the system. Discussion about the results obtained during the implementation case are presented and the potential of using the information generated from the system as a managing tool is commented.

  20. ABC3 Consensus: Assessment by a German Group of Experts.

    Science.gov (United States)

    Thomssen, Christoph; Augustin, Doris; Ettl, Johannes; Haidinger, Renate; Lück, Hans-Joachim; Lüftner, Diana; Marmé, Frederik; Marschner, Norbert; Müller, Lothar; Overkamp, Friedrich; Ruckhäberle, Eugen; Thill, Marc; Untch, Michael; Wuerstlein, Rachel; Harbeck, Nadia

    2016-02-01

    The Advanced Breast Cancer Third International Consensus Conference on the diagnosis and treatment of advanced breast cancer took place in Lisbon, Portugal, on November 5-7, 2015. This year's conference (ABC3) was focused on the treatment of metastatic breast cancer (stage IV), as it was 4 years ago at the first consensus meeting (ABC1). A matter of particular interest was the patients' perspective. Thus, patient-relevant issues were addressed by the consensus discussions, such as those on treatment goals, quality of life, care of long-term survivors ('survivorship issues'), and coping with disease-related symptoms and the side effects of treatment. Further important issues on the agenda were the use of standardized instruments for the assessment of individual treatment success ('patient-reported outcome measures') and the evaluation of the benefit of novel drugs (e.g. the European Society for Medical Oncology (ESMO) Magnitude of Clinical Benefit Scale). Diagnosis and treatment of inoperable locally advanced breast cancer had already been discussed 2 years earlier at the ABC2 Consensus and were not dealt with in the framework of this year's ABC3 Consensus. With regard to country-specific peculiarities, which unavoidably found their way into the ABC Consensus, a working group of German breast cancer experts commented on the voting results of the ABC panelists. As for the past consensus, the group specially considered the German guidelines for the diagnosis and treatment of breast cancer (AGO (Gyneco-Oncology Working Group), S3, DGHO (German Society of Hematology and Medical Oncology)) in order to adapt the ABC3 consensus for everyday therapy in Germany.