Papadopoulou B, Roy G, Dey S, Rosen B P, Ouellette M
Service d'Infectiologie du Centre de Recherche du CHUL, Québec, Canada.
J Biol Chem. 1994 Apr 22;269(16):11980-6.
Oxyanions in the form of pentavalent antimony compounds are currently the drug of choice for treating leishmaniasis. Leishmania mutants resistant to high concentrations of the oxyanion arsenite were obtained in a stepwise selection procedure. Amplification of the H locus P-glycoprotein-related gene ltpgpA, as part of extrachromosomal circles, is a frequent event in arsenite-resistant cells, but was observed only in cells resistant to high concentrations of the metalloid salts. Revertants grown in the absence of the drug lost their ltpgpA-containing amplicon and part of their resistance. The results of previous transfection experiments in Leishmania had suggested that ltpgpA is only involved in low level resistance to arsenite and antimonite. The results of this study using transfection of ltpgpA alleles isolated from arsenite-resistant mutants or of whole circular amplicons containing ltpgpA demonstrate clearly that this P-glycoprotein-related gene is involved in low level resistance to oxyanions, including the pentavalent antimony-containing drug Pentostam. By site-directed mutagenesis, the LtpgpA protein was shown to require an intact nucleotide-binding site to confer arsenite resistance. Under the experimental conditions used, decreased accumulation of the 73AsO2- drug was not observed in the ltpgpA transfectants. One possibility is that LtpgpA-mediated arsenite resistance could result from sequestration of the toxic anion in an intracellular compartment. The results indicate that despite the fact that ltpgpA amplification is a frequent event in oxyanion-resistant mutants, it contributes only slightly to the overall resistance.
五价锑化合物形式的含氧阴离子目前是治疗利什曼病的首选药物。通过逐步筛选程序获得了对高浓度含氧阴离子亚砷酸盐具有抗性的利什曼原虫突变体。作为染色体外环状物一部分的H位点P - 糖蛋白相关基因ltpgpA的扩增，在抗亚砷酸盐细胞中是常见事件，但仅在对高浓度类金属盐具有抗性的细胞中观察到。在无药物条件下生长的回复突变体失去了其含ltpgpA的扩增子及部分抗性。先前在利什曼原虫中的转染实验结果表明，ltpgpA仅参与对亚砷酸盐和亚锑酸盐的低水平抗性。本研究使用从抗亚砷酸盐突变体中分离的ltpgpA等位基因或包含ltpgpA的完整环状扩增子进行转染的结果清楚地表明，这个P - 糖蛋白相关基因参与对含氧阴离子的低水平抗性，包括含五价锑的药物喷他脒。通过定点诱变表明，LtpgpA蛋白需要完整的核苷酸结合位点来赋予亚砷酸盐抗性。在所使用的实验条件下，在ltpgpA转染体中未观察到73AsO2-药物积累的减少。一种可能性是LtpgpA介导的亚砷酸盐抗性可能是由于在细胞内区室中隔离有毒阴离子所致。结果表明，尽管ltpgpA扩增在抗含氧阴离子突变体中是常见事件，但它对总体抗性的贡献仅微乎其微。
