Gene Expression Profiling and Molecular Characterization of Antimony Resistance in Leishmania amazonensis 英文参考文献.docVIP

GeneExpressionProfilingandMolecular CharacterizationofAntimonyResistanceinLeishmania amazonensis RubensL.doMonte-Neto1,2,AdrianoC.Coelho2,Fre′de′ricRaymond2,3,DanielleLe′gare′2, JacquesCorbeil2,3,MariaN.Melo4,Fre′de′ricFre′zard1,MarcOuellette2* 1DepartamentodeFisiologiaeBiof?′sica,InstitutodeCie?nciasBiolo′gicas,UniversidadeFederaldeMinasGerais,BeloHorizonte,MinasGerais,Brasil,2CentredeRecherche en Infectiologie du Centre de Recherche du CHUL and De′partement de Microbiologie, Immunologie et Infectiologie, Universite′ Laval, Que′bec, Que′bec, Canada, 3De′partement de Me′decine Mole′culaire Faculte′ de Me′decine, Universite′ Laval, Que′bec, Que′bec, Canada, 4Departamento de Parasitologia, Instituto de Cie?ncias Biolo′gicas,UniversidadeFederaldeMinasGerais,BeloHorizonte,MinasGerais,Brasil Abstract Background: Drug resistance is a major problem in leishmaniasis chemotherapy. RNA expression profiling using DNA microarraysisasuitableapproachtostudysimultaneouseventsleadingtoadrug-resistancephenotype.Genomicanalysis hasbeenperformedprimarilywithOldWorldLeishmaniaspeciesandhereweinvestigatemolecularalterationsinantimony resistanceintheNewWorldspeciesL.amazonensis. Methods/PrincipalFindings:WeselectedpopulationsofL.amazonensispromastigotesforresistancetoantimonybystep- wise drug pressure. Gene expression of highly resistant mutants was studied using DNA microarrays. RNA expression profilingofantimony-resistantL.amazonensisrevealedtheoverexpressionofgenesinvolvedindrugresistanceincluding the ABC transporter MRPA and several genes related to thiol metabolism. The MRPA overexpression was validated by quantitative real-time RT-PCR and further analysis revealed that this increased expression was correlated to gene amplificationaspartofextrachromosomallinearampliconsinsomemutantsandaspartofsupernumerarychromosomesin other mutants. The expression of several other genes encoding hypothetical proteins but also nucleobase and glucose transporterencodinggeneswerefoundtobemodulated.