Deconvoluting heme biosynthesis to target blood-stage malaria parasites.
Deconvoluting heme biosynthesis to target blood-stage malaria parasites.
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DOI：
10.7554/elife.09143
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发表时间：
2015-07-14
期刊：
影响因子：
7.7
通讯作者：
Goldberg DE
中科院分区：
文献类型：
--
作者：
Sigala PA;Crowley JR;Henderson JP;Goldberg DE
血红素代谢是疟疾寄生虫恶性疟原虫血液阶段感染的核心。寄生虫保留血红素生物合成途径，但在感染富含血红素的红细胞期间不需要其活性，在那里它们可以吸收宿主血红素以满足代谢需要。然而，血红素生物合成寄生虫感染的红细胞可以有力地刺激外源性5-氨基乙酰丙酸（ALA），导致积累的光毒性中间体原卟啉IX（PPIX）。在这里，我们使用光动力学成像，质谱，寄生虫基因破坏，和化学探针揭示疟原虫感染的红细胞的细胞质中的残留宿主酶有助于ALA刺激血红素的生物合成和ALA摄取依赖于寄生虫建立的渗透性途径。我们发现，PPIX在感染的红细胞中的积累可以利用基于鲁米诺的化学发光和低剂量青蒿素的组合刺激来光活化PPIX以产生细胞毒性活性氧来进行抗疟化疗。这种光动力学策略具有利用对赋予抗性的突变难治的宿主酶的优点。DOI：http://dx.doi.org/10.7554/eLife.09143.001疟疾是一种毁灭性的传染病，是由单细胞疟原虫引起的，疟原虫可以生活在红细胞内。这些寄生虫的几种重要蛋白质需要一种叫做血红素的小分子才能发挥作用。寄生虫有酶，通过一系列中间步骤制造血红素。然而，目前还不清楚这种酶的“途径”对寄生虫有多重要，以及这种途径是否可以被药物靶向治疗疟疾。现在，Sigala等人已经使用了一系列遗传和生物化学方法来更好地了解疟原虫感染的红细胞中血红素分子的产生。首先，删除了几个编码用于制造血红素分子的酶的寄生虫基因。出乎意料的是，这些基因缺失并没有影响受感染的血细胞制造血红素的能力。这一结果表明，寄生虫在血液中生长时并不使用自己的途径来产生血红素。Sigala等人随后发现，人类参与制造血红素的酶仍然活跃，其中大部分也存在于受感染的红细胞中。这些人类酶提供了一个平行的途径，可以连接到最终的寄生虫酶，以产生血红素。进一步的实验表明，通过为该途径提供用于制造血红素的构建块之一，可以强烈刺激人类酶的活性。这种刺激导致了一种叫做PPIX的中间分子的形成。当这种中间分子暴露在光下时，它可以杀死细胞，这种特性被称为“光毒性”。Sigala等人表明，用一种新的无毒发光化学物质组合治疗感染的红细胞可以激活血液中的PPIX，并可以选择性地杀死疟疾寄生虫，同时保持未感染细胞的完整性。这些发现提出了一种新的治疗方法，可以有效地对抗血液阶段的疟疾。此外，寄生虫将无法轻易突变以避免这种治疗的影响，因为它依赖于已经制造的人类蛋白质。现在需要进一步的工作来优化可以提供这种治疗的药物的剂量和组合。DOI：http://dx.doi.org/10.7554/eLife.09143.002
Heme metabolism is central to blood-stage infection by the malaria parasite Plasmodium falciparum. Parasites retain a heme biosynthesis pathway but do not require its activity during infection of heme-rich erythrocytes, where they can scavenge host heme to meet metabolic needs. Nevertheless, heme biosynthesis in parasite-infected erythrocytes can be potently stimulated by exogenous 5-aminolevulinic acid (ALA), resulting in accumulation of the phototoxic intermediate protoporphyrin IX (PPIX). Here we use photodynamic imaging, mass spectrometry, parasite gene disruption, and chemical probes to reveal that vestigial host enzymes in the cytoplasm of Plasmodium-infected erythrocytes contribute to ALA-stimulated heme biosynthesis and that ALA uptake depends on parasite-established permeability pathways. We show that PPIX accumulation in infected erythrocytes can be harnessed for antimalarial chemotherapy using luminol-based chemiluminescence and combinatorial stimulation by low-dose artemisinin to photoactivate PPIX to produce cytotoxic reactive oxygen. This photodynamic strategy has the advantage of exploiting host enzymes refractory to resistance-conferring mutations. DOI: http://dx.doi.org/10.7554/eLife.09143.001 Malaria is a devastating infectious disease that is caused by single-celled parasites called Plasmodium that can live inside red blood cells. Several important proteins from these parasites require a small molecule called heme in order to work. The parasites have enzymes that make heme via a series of intermediate steps. However, it remains unclear exactly how important this ‘pathway’ of enzymes is for the parasite, and whether this pathway could be targeted by drugs to treat malaria. Now Sigala et al. have used a range of genetic and biochemical approaches to better understand the production of heme molecules in Plasmodium-infected red blood cells. First, several parasite genes that encode the enzymes used to make heme molecules were deleted. Unexpectedly, these gene deletions did not affect the ability of the infected blood cells to make heme. This result suggested that the parasites do not use their own pathway to produce heme while they are growing in the bloodstream. Sigala et al. then showed that human enzymes involved in making heme, most of which are also found within the infected red blood cells, are still active. These human enzymes provide a parallel pathway that can link up with the final parasite enzyme to generate heme. Further experiments revealed that the activity of the human enzymes could be strongly stimulated by providing the pathway with one of the building blocks used to make heme. This stimulation led to the build-up of an intermediate molecule called PPIX. This intermediate molecule can kill cells when it is exposed to light—a property that is called ‘phototoxicity’. Sigala et al. showed that treating infected red blood cells with a new combination of non-toxic chemicals that emit light can activate PPIX in the bloodstream and can selectively kill the malaria parasites while leaving uninfected cells intact. These findings suggest a new treatment that could be effective against blood-stage malaria. Furthermore, the parasite will be unable to easily mutate to avoid the effects of this treatment because it relies on human proteins that are already made. Future work is now needed to optimize the dosage and the combination of drugs that could provide such a treatment. DOI: http://dx.doi.org/10.7554/eLife.09143.002