Cloning, functional analysis and post-transcriptional regulation of a type II DNA topoisomerase from Leishmania infantum.: A new potential target for anti-parasite drugs
Cloning, functional analysis and post-transcriptional regulation of a type II DNA topoisomerase from Leishmania infantum.: A new potential target for anti-parasite drugs
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DOI：
10.1093/nar/gkg671
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发表时间：
2003-08-15
影响因子：
14.9
通讯作者：
Larraga, V
中科院分区：
文献类型：
--
作者：
Hanke, T;Ramiro, MJ;Larraga, V
我们从婴儿利什曼原虫（一种引起人类疾病的寄生虫原虫）中鉴定出一种 II 型拓扑异构酶。这种名为 Li topo II 的蛋白质位于动质体中，具有可变的 C 末端。编码 Li-TOP2 的克隆基因补偿了酿酒酵母拓扑 II 缺陷突变体的缓慢生长，从而在酵母中产生具有催化活性的 DNA 拓扑异构酶。对 Li-TOP2 基因的特定 mRNA 水平的分析显示，同步化细胞中的整个寄生虫细胞周期以及不同形式的寄生虫之间存在差异。因此，该酶在寄生虫的高感染性细胞内形式（无鞭毛体）中比在细胞外前鞭毛体形式中具有更高水平的 mRNA 表达，这表明与原虫的不同发育和感染阶段有关。此外，蛋白质印迹分析显示，增殖型和非增殖型婴儿乳杆菌前鞭毛体之间的蛋白质表达存在差异，其 mRNA 水平相似。这表明可能的转录后调节机制。数据表明，Li topo II 参与 DNA 串联，并且可能参与婴儿 L.infantum 细胞内形式增殖的初始阶段，L.infantum 是一种寄生虫，在细胞内感染的最初时刻必须增殖到宿主巨噬细胞中才能在其恶劣环境中生存。
We identified a type II topoisomerase enzyme from Leishmania infantum, a parasite protozoon causing disease in humans. This protein, named Li topo II, which displays a variable C-terminal end, is located in the kinetoplast. The cloned gene encoding Li-TOP2 compensates for the slow growth of topo II-deficient mutants of Saccharomyces cerevisiae, resulting in a catalytically active DNA topoisomerase in yeast. Analysis of the specific mRNA levels of the Li-TOP2 gene showed variations throughout the parasite cell cycle in synchronized cells as well as between the distinct forms of the parasite. Thus, the enzyme had higher levels of mRNA expression in the highly infective intracellular form of the parasite, the amastigote, than in the extracellular promastigote form, suggesting a relation with the distinct developmental and infectious phases of the protozoon. In addition, western blot analysis showed differences in protein expression between the proliferative and non-proliferative forms of L.infantum promastigotes, which displayed similar levels of mRNA. This indicated possible post-transcriptional regulation mechanisms. The data suggest that Li topo II has a part in DNA decatenation and probably at the initial stages of proliferation in the intracellular form of L.infantum, a parasite that has to proliferate into the host macrophage to survive its hostile environment in its first moments of intracellular infection.