Genome-wide RNAi screens in bloodstream form trypanosomes identify drug transporters

被引：161

作者：

Burkard, Gabriela Schumann ^{[1
]}

Jutzi, Pascal ^{[1
]}

Roditi, Isabel ^{[1
]}

机构：

[1] Univ Bern, Inst Cell Biol, CH-3012 Bern, Switzerland

来源：

MOLECULAR AND BIOCHEMICAL PARASITOLOGY | 2011年 / 175卷 / 01期

关键词：

Trypanosoma brucci; Melarsoprol; Eflornithine; Amino acid; Transporter; Drug resistance; INDUCIBLE EXPRESSION SYSTEM; BRUCEI-RHODESIENSE; GLYCOPROTEIN; GLYCOLYSIS; LIBRARY;

D O I：

10.1016/j.molbiopara.2010.09.002

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

An inducible RNA interference (RNAi) library, consisting of a pool of independent stable transformants with 9-fold genome coverage, was constructed in bloodstream form Trypanosoma brucei using an improved transfection protocol. RNAi induction and selection of resistant parasites was performed in the presence of melarsoprol or eflornithine. The former led to the isolation of the adenosine transporter TbAT1, which is known to be involved in melarsoprol uptake, while the latter identified an amino acid transporter, AAT6. Knockdown of AAT6 reduced mRNA levels to 30-35% in independent clones and increased resistance to eflornithine >5-fold. Genome-wide screens with this library allow an unbiased approach to gene discovery, are extremely rapid and do not exclude essential genes. (C) 2010 Elsevier B.V. All rights reserved.

引用

页码：91 / 94

页数：4

共 12 条

[1] Highly efficient stable transformation of bloodstream forms of Trypanosoma brucei [J].

Burkard, Gabriela ;

Fragoso, Cristina M. ;

Roditi, Isabel .

MOLECULAR AND BIOCHEMICAL PARASITOLOGY, 2007, 153 (02) :220-223

[2] The adenosine analog tubercidin inhibits glycolysis in Trypanosoma brucei as revealed by an RNA interference library [J].

Drew, ME ;

Morris, JC ;

Wang, ZF ;

Wells, L ;

Sanchez, M ;

Landfear, SM ;

Englund, PT .

JOURNAL OF BIOLOGICAL CHEMISTRY, 2003, 278 (47) :46596-46600

[3] High-throughput RNAi screening to dissect cellular pathways: A how-to guide [J].

Falschlehner, Christina ;

Steinbrink, Sandra ;

Erdmann, Gerrit ;

Boutros, Michael .

BIOTECHNOLOGY JOURNAL, 2010, 5 (04) :368-376

[4] N-myristoyltransferase inhibitors as new leads to treat sleeping sickness [J].

Frearson, Julie A. ;

Brand, Stephen ;

McElroy, Stuart P. ;

Cleghorn, Laura A. T. ;

Smid, Ondrej ;

Stojanovski, Laste ;

Price, Helen P. ;

Guther, M. Lucia S. ;

Torrie, Leah S. ;

Robinson, David A. ;

Hallyburton, Irene ;

Mpamhanga, Chidochangu P. ;

Brannigan, James A. ;

Wilkinson, Anthony J. ;

Hodgkinson, Michael ;

Hui, Raymond ;

Qiu, Wei ;

Raimi, Olawale G. ;

van Aalten, Daan M. F. ;

Brenk, Ruth ;

Gilbert, Ian H. ;

Read, Kevin D. ;

Fairlamb, Alan H. ;

Ferguson, Michael A. J. ;

Smith, Deborah F. ;

Wyatt, Paul G. .

NATURE, 2010, 464 (7289) :728-U100

[5] Alterations in ornithine decarboxylase characteristics account for tolerance of Trypanosoma brucei rhodesiense to D,L-alpha-difluoromethylornithine [J].

Iten, M ;

Mett, H ;

Evans, A ;

Enyaru, JCK ;

Brun, R ;

Kaminsky, R .

ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, 1997, 41 (09) :1922-1925

[6] A nucleoside transporter from Trypanosoma brucei involved in drug resistance [J].

Mäser, P ;

Sütterlin, C ;

Kralli, A ;

Kaminsky, R .

SCIENCE, 1999, 285 (5425) :242-244

[7] Glycolysis modulates trypanosome glycoprotein expression as revealed by an RNAi library [J].

Morris, JC ;

Wang, ZF ;

Drew, ME ;

Englund, PT .

EMBO JOURNAL, 2002, 21 (17) :4429-4438

[8] The Alamar Blue(R) assay to determine drug sensitivity of African trypanosomes (T-b-rhodesiense and T-b-gambiense) in vitro [J].

Raz, B ;

Iten, M ;

GretherBuhler, Y ;

Kaminsky, R ;

Brun, R .

ACTA TROPICA, 1997, 68 (02) :139-147

[9] PROCYCLIN GENE-EXPRESSION AND LOSS OF THE VARIANT SURFACE GLYCOPROTEIN DURING DIFFERENTIATION OF TRYPANOSOMA-BRUCEI [J].

RODITI, I ;

SCHWARZ, H ;

PEARSON, TW ;

BEECROFT, RP ;

LIU, MK ;

RICHARDSON, JP ;

BUHRING, HJ ;

PLEISS, J ;

BULOW, R ;

WILLIAMS, RO ;

OVERATH, P .

JOURNAL OF CELL BIOLOGY, 1989, 108 (02) :737-746

[10] Mitochondrial membrane potential-based genome-wide RNAi screen of Trypanosoma brucei [J].

Verner, Zdenek ;

Paris, Zdenek ;

Lukes, Julius .

PARASITOLOGY RESEARCH, 2010, 106 (05) :1241-1244

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