G-quadruplex biology in the human malaria parasite Plasmodium falciparum

这项拟议的研究涉及最重要的人类疟疾寄生虫——恶性疟原虫。疟疾是世界上最使人衰弱的传染病之一，每年造成50多万人死亡，影响数亿人。大多数死亡发生在撒哈拉以南非洲的幼儿中，但成年人也可能终生患疟疾，在疟疾流行国家降低生活质量，阻碍经济发展。缺乏有效的疫苗和耐药寄生虫的出现意味着现在迫切需要进行研究，以便更好地了解疟疾寄生虫，从而找到针对这种疾病的新疫苗靶点和治疗战略。疟疾寄生虫通过感染红细胞引起疾病。它在这些细胞内繁殖，用一种叫做pfemp1的蛋白质修饰细胞表面，这种蛋白质与血管壁结合。这对寄生虫的生存至关重要，因为它可以将感染的细胞从循环血液中移除，并保护它们不通过可能识别并摧毁它们的脾脏。它还会导致疾病，严重疟疾尤其与受感染细胞在脑血管和胎盘中的积聚有关。因此，疟疾生物学家非常感兴趣的是了解控制这些粘附性PfEMP1蛋白表达的机制。并非所有疟疾寄生虫都统一表达pfemp1：相反，个体寄生虫经常在不同的变体之间切换。这使他们能够领先于免疫系统，并维持慢性感染数月甚至数年。寄生虫有一个大的、可变的“var”基因家族，用于不同的PfEMP1蛋白，它们通过所谓的“表观遗传开关”改变这些基因的表达。此外，变异基因很容易重组产生新的变异，因此每种寄生虫菌株——在流行地区有数百种——都有独特的可能的表面蛋白质。这就是为什么人类对反复感染疟疾的免疫力发展缓慢的原因之一：每一种新的寄生虫菌株在免疫系统看来都是不同的，因此人们可能在一生中反复感染。了解并最终干扰var基因的表达、切换和重组，从而干扰PfEMP1蛋白的变异表达，可能是更有效地免疫控制疟疾的关键。因此，本研究的重点是寄生虫在var基因之间切换和产生新变体的一种新的生物学机制。我们最近的工作表明，一种不寻常的DNA结构，称为g -四重体，集中在var基因周围，似乎影响了这两个过程。为了进一步研究这一点，我们现在建议在整个疟原虫基因组中绘制g -四联体，包括DNA和信使分子RNA。我们将使用一系列尖端的全基因组技术来做到这一点，然后检查当解除它们的酶被移除时，结构的分布是否会发生变化。这些研究将导致更好地理解var基因动态背后的机制，并可能最终为抗击疟疾的新策略提供信息，因为var基因——以及它们编码的粘附蛋白——是疟疾疾病的核心。研究结果将发表在开放获取的科学期刊上，并在国际会议上发表。它们将通过适当网站上的摘要以及杂志和/或网上的科学文章向公众传播。只要疟疾寄生虫继续造成巨大的人类疾病负担，这类工作就仍然至关重要。

The proposed research concerns the most important human malaria parasite, Plasmodium falciparum. Malaria is one of the world's most debilitating infectious diseases, killing over half a million people every year and affecting several hundred million. Most of the deaths occur in young children in sub-Saharan Africa, but adults can also suffer from malaria throughout their lives, reducing quality of life and retarding economic development in endemic countries. The lack of an effective vaccine and the emergence of drug-resistant parasites mean that there is now an urgent need for research leading to a better understanding of the malaria parasite, and hence to new vaccine targets and treatment strategies for this disease.The malaria parasite causes illness via the infection of red blood cells. It multiplies inside these cells and modifies their surfaces with proteins called PfEMP1s that bind to the walls of blood vessels. This is crucial for parasite survival as it removes infected cells from the circulating blood and protects them from passing through the spleen, which might recognize and destroy them. It also contributes to disease, with severe malaria being particularly associated with the accumulation of infected cells in vessels of the brain and placenta. It is therefore of great interest to malaria biologists to understand the mechanisms that control the expression of these adhesive PfEMP1 proteins. PfEMP1s are not expressed uniformly by all malaria parasites: instead, individual parasites regularly switch between different variants. This allows them to stay ahead of the immune system and sustain a chronic infection for months or even years. The parasites have a large, variable family of 'var' genes for different PfEMP1 proteins and they vary the expression of these genes by so-called 'epigenetic switching'. Furthermore, var genes recombine very readily to generate new variants, so each parasite strain - of which there are many hundreds circulating in endemic areas - has a unique repertoire of possible surface proteins. This is one reason why immunity to repeated malaria infections is slow to develop in humans: every new parasite strain looks different to the immune system, so people can be re-infected repeatedly throughout their lives.Understanding and ultimately interfering with the expression, switching and recombination of var genes, and thus the variant expression of PfEMP1 proteins, could be a key to more effective immune control of malaria. Therefore, this research focuses on a new biological mechanism that the parasite may use for switching between var genes and for generating new variants. Our recent work has showed that an unusual DNA structure called a G-quadruplex that is concentrated around var genes seems to affect both these processes. To investigate this further, we now propose to map the G-quadruplexes throughout the Plasmodium genome, both in DNA and also in the messenger molecule, RNA. We will use a range of cutting-edge genome-wide technologies to do this, and will then check whether the distribution of the structures changes when the enzymes that unwind them are removed.These studies will lead to a better understanding of the mechanisms underlying var gene dynamics, and may ultimately inform new strategies to combat malaria, since var genes - and the adhesive proteins that they encode - are central to malarial disease. The outcomes of the research will be published in open-access scientific journals and presented at international conferences. They will be communicated to the general public via summaries on appropriate websites and via science writing in magazines and/or online. Work such as this remains vital as long as the malaria parasite continues to cause an immense burden of human disease.