Effect of polyglutamylation of methotrexate on its accumulation and the development of resistance in the protozoan parasite Leishmania
Effect of polyglutamylation of methotrexate on its accumulation and the development of resistance in the protozoan parasite Leishmania
复制标题
DOI：
10.1016/s0006-2952(03)00417-9
复制
发表时间：
2003-09-15
影响因子：
5.8
通讯作者：
Ouellette, M
中科院分区：
文献类型：
--
作者：
El Fadili, A;Richard, D;Ouellette, M
在大多数生物体中，叶酸是多谷氨酸化的，而在癌细胞中，叶酸和抗叶酸氨甲喋呤(MTX)的多谷氨酰化是MTX易感性的重要决定因素。最近在利什曼原虫中发现了负责叶酸多谷氨酰化的叶基多谷氨酸合成酶(FPGS)。我们在这里表明，在塔伦托利什曼原虫中，MTX是多谷氨酸化的，并且三聚氰酸酯是主要的形式。转导FPGS基因的利什曼原虫细胞中，MTX的谷氨酸链长度显著增加，而在一个FPGS等位基因被破坏的细胞中，MTX的谷氨酸链长度明显减少。对FPGS基因表达的调节对MTX的敏感性也有深远的影响，这种影响被发现依赖于介质中的叶酸浓度。在富含叶酸的SDM-79培养液中，过量表达的FPGS对MTX产生抗性，而在叶酸含量低得多的M-199培养液中，转染者对MTX更为敏感。在低叶酸培养条件下，一个FPGS等位基因被破坏的细胞对MTX的抗性更强。FPGS表达的调节影响了叶酸和MTX的短期和长期积累，显示出FPGS单倍体突变体的积累显著减少。这种不同的蓄积是通过药物流入细胞的减少来调节的。最后，对MTX抗性的利什曼原虫突变株的分析表明，MTX上存在较短的谷氨酸链与MTX抗性有关。(C)2003 Elsevier Inc.保留所有权利。
Folates are polyglutamylated in most organisms, and in cancer cells the polyglutamylation of folates and of the antifolate methotrexate (MTX) is an important determinant of MTX susceptibility. The folylpolyglutamate synthetase (FPGS) responsible for polyglutamylation of folates was recently characterized in the parasite Leishmania. We show here that MTX is polyglutamylated in Leishmania tarentolae and that triglutamates are the predominant form. The glutamate chain length of MTX increases significantly in Leishmania cells transfected with the FPGS gene and decreases in cells with one FPGS allele disrupted. Modulation in the expression of the FPGS gene also has a profound effect on MTX susceptibility and this effect was found to be dependent on the folate concentration of the medium. In the folate-rich medium SDM-79, overexpression of FPGS will confer MTX resistance while in M-199 medium, which has much less folates, FPGS transfectants are more sensitive to MTX. Cells with one allele of FPGS disrupted are more resistant to MTX in low folate medium. The modulation of FPGS expression affects both the short-term and long-term accumulation of folate and MTX, showing a marked decrease in accumulation in the FPGS haploid mutant. This differential accumulation was mediated by decreased influx of the drug into the cell. Finally, the analysis of MTX-resistant Leishmania mutants indicated that the presence of shorter glutamate chains on MTX is correlated with MTX resistance. (C) 2003 Elsevier Inc. All rights reserved.