A genetically hard-wired metabolic transcriptome in Plasmodium falciparum fails to mount protective responses to lethal antifolates.
A genetically hard-wired metabolic transcriptome in Plasmodium falciparum fails to mount protective responses to lethal antifolates.
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DOI：
10.1371/journal.ppat.1000214
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发表时间：
2008-11
期刊：
影响因子：
6.7
通讯作者：
Rathod PK
中科院分区：
文献类型：
--
作者：
Ganesan K;Ponmee N;Jiang L;Fowble JW;White J;Kamchonwongpaisan S;Yuthavong Y;Wilairat P;Rathod PK
恶性疟原虫的基因组序列允许全球分析抗疟药物的药物反应。了解DNA微阵列如何用于研究疟疾寄生虫的药物作用是很有意义的。在一项大型的严格控制的研究中，涉及123个来自同基因药物敏感和耐药寄生虫的cDNA之间的微阵列杂交，致命的抗叶酸剂（WR 99210）未能过度产生遗传学证明的主要靶点二氢叶酸还原酶-胸苷酸合酶（DHFR-TS）的RNA。这种转录刚性延续到代谢相关的RNA编码叶酸和嘧啶生物合成，以及寄生虫基因组的其余部分。在初始药物暴露后24 h内，没有基因可再现地上调超过2倍，即使克隆活力在6 h内下降了50%。我们预测并表明，虽然寄生虫不会在真实的时间内对抗叶酸剂产生保护性转录反应，但转染了人DHFR基因并适应长期WR 99210暴露的恶性疟原虫细胞会调整硬连线转录组本身，使其在药物存在下茁壮成长。一个系统范围内的能力，改变RNA水平，以响应特定的代谢扰动可能会导致恶性疟原虫致命的抗代谢物的选择性脆弱性。此外，这种调节影响了DNA微阵列如何用于了解抗代谢药的作用模式。基因调控的传统知识主要是从非致病性模式生物（例如大肠杆菌）中学到的。大肠杆菌、酵母和小鼠的研究表明，代谢途径中的RNA在很大程度上受到DNA结合转录因子的调节，这些转录因子对细胞代谢需求有反应。我们证明，疟疾引起的恶性疟原虫寄生虫，在致命的药物压力下，从抗叶酸剂与已知的作用机制，是无法大的可重复的变化，在RNA水平的目标途径，或在整个基因组的任何其他基因。在垂死的寄生虫中可以检测到小RNA的变化，可能是干扰途径的信息。此外，当控制RNA水平的硬连线程序本身被改变时，可以看到显著的RNA变化。我们的数据正式证明，疟原虫中间代谢的RNA水平在很大程度上是预先确定的。我们建议，作为一个寄生虫与复杂的生命周期旅行从一个很大程度上可预测的细胞内生化环境到另一个，这样的硬布线可能足以管理转录水平的中间代谢，而不采用感官功能。当重要的靶途径在寄生虫中是刚性的但在宿主细胞中动态调节时，这种全系统的宿主-寄生虫基因调控差异可能会产生意想不到的药理学机会。
Genome sequences of Plasmodium falciparum allow for global analysis of drug responses to antimalarial agents. It was of interest to learn how DNA microarrays may be used to study drug action in malaria parasites. In one large, tightly controlled study involving 123 microarray hybridizations between cDNA from isogenic drug-sensitive and drug-resistant parasites, a lethal antifolate (WR99210) failed to over-produce RNA for the genetically proven principal target, dihydrofolate reductase-thymidylate synthase (DHFR-TS). This transcriptional rigidity carried over to metabolically related RNA encoding folate and pyrimidine biosynthesis, as well as to the rest of the parasite genome. No genes were reproducibly up-regulated by more than 2-fold until 24 h after initial drug exposure, even though clonal viability decreased by 50% within 6 h. We predicted and showed that while the parasites do not mount protective transcriptional responses to antifolates in real time, P. falciparum cells transfected with human DHFR gene, and adapted to long-term WR99210 exposure, adjusted the hard-wired transcriptome itself to thrive in the presence of the drug. A system-wide incapacity for changing RNA levels in response to specific metabolic perturbations may contribute to selective vulnerabilities of Plasmodium falciparum to lethal antimetabolites. In addition, such regulation affects how DNA microarrays are used to understand the mode of action of antimetabolites. Traditional knowledge of gene regulation, learned largely from non-pathogenic model organisms such as E. coli, yeast, and mice, suggests that RNA for metabolic pathways are regulated in large part by DNA-binding transcriptional factors that are responsive to cellular metabolic needs. We demonstrate that the malaria-causing Plasmodium falciparum parasites, under lethal drug pressure from an antifolate with a known mechanism of action, are incapable of large reproducible changes in RNA levels for the target pathways, or for any other gene throughout the genome. Small RNA changes, possibly informative of perturbed pathways, can be detected in dying parasites. In addition, significant RNA changes are seen when the hard-wired program, governing RNA levels, itself is altered. Our data formally proves that RNA levels for intermediary metabolism in malaria parasites are largely predetermined. We propose that as a parasite with a complex life cycle travels from one largely predictable intracellular biochemical environment to another, such hard-wiring may be sufficient to manage transcript levels for intermediary metabolism without employing sensory functions. Such a system-wide host–parasite difference in gene regulation may create unexpected pharmacological opportunities when important target pathways are rigid in the parasite but dynamically regulated in host cells.
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