Tetracycline analogue-regulated transgene expression in Plasmodium falciparum blood stages using Toxoplasma gondii transactivators
Tetracycline analogue-regulated transgene expression in Plasmodium falciparum blood stages using Toxoplasma gondii transactivators
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DOI：
10.1073/pnas.0500112102
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发表时间：
2005-02-22
影响因子：
11.1
通讯作者：
Crabb, BS
中科院分区：
文献类型：
--
作者：
Meissner, M;Krejany, E;Crabb, BS
基因操作彻底改变了对顶角丛寄生虫恶性疟原虫的研究，恶性疟原虫是疟疾最重要的病原体。然而，迄今为止，还没有建立允许对血液阶段生长有害的修饰的技术，例如破坏必需基因或显性负转基因的表达。最近建立了一个筛选功能的反式激活因子在相关的寄生虫弓形虫，促使我们确定反式激活因子在T。弓形虫，并检查其在恶性疟原虫中的功能。四环素应答性最小启动子基于表征的恶性疟原虫钙调蛋白启动子产生，并用于评估恶性疟原虫中的反式激活因子。我们证明了T.弓形虫在恶性疟原虫中也有功能。通过使用四环素类似物脱水四环素，在恶性疟原虫中实现了有效的阶段特异性基因调控。这种可调控的表达技术对于研究恶性疟原虫血液阶段的必需基因功能具有明显的潜力。另一方面，所鉴定的反式激活因子在哺乳动物细胞中不起作用，这与顶复门寄生虫及其人类宿主之间基因调控机制的根本差异一致。
Genetic manipulation has revolutionized research in the Apicornplexan parasite Plasmodium falciparum, the most important causative agent of malaria. However, to date no techniques have been established that allow modifications that are deleterious to blood-stage growth, such as the disruption of essential genes or the expression of dominant-negative transgenes. The recent establishment of a screen for functional transactivators in the related parasite Toxoplasma gondii prompted us to identify transactivators in T. gondii and to examine their functionality in A falciparum. Tetracycline-responsive minimal promoters were generated based on the characterized P. falciparum calmodulin promoter and used to assess transactivators in A falciparum. We demonstrate that artificial tetracycline-regulated transactivators isolated in T. gondii are also functional in P. falciparum. By using the tetracycline analogue anhydrotetracycline, efficient, stage-specific gene regulation was achieved in P. falciparum. This regulatable expression technology has clear potential for the study of essential gene function in A falciparum blood stages. On the other hand, the identified transactivators are not functional in mammalian cells, consistent with the fundamental differences in the mechanism of gene regulation between Apicomplexan parasites and their human hosts.