Copy Number Variation in Malaria Parasites

描述（由申请人提供）：基因拷贝数变异（CNV）在真核基因组中普遍存在，并且由此产生的剂量变化在确定表型中起重要作用。我们的目标是定义的程度，进化动力学和功能的CNV在疟疾寄生虫基因组（恶性疟原虫）。为了实现这一点，我们将使用比较基因组杂交（CGH），它提供了一个有效的工具，用于测定CNV和单核苷酸多态性（SNP），是特别有效的单倍体基因组。初步数据揭示了恶性疟原虫基因组中丰富的CNV（缺失和扩增），并证明了SNP的准确预测。该项目将分为三个部分。首先，我们将确定所有的CNV和目录侧翼SNP在世界范围内的寄生虫收集。抽样人群将包括来自泰国、巴布亚新几内亚、哥伦比亚和冈比亚的新分离的单克隆感染者。我们将测量基因组重排的大小、基因含量、排列和群体频率。我们的第二个目标将是评估基因组区域适应性进化的证据，显示CNV突出可能受到最近进化压力（如药物选择）影响的基因组片段。我们将通过研究侧翼SNP多态性的连锁不平衡模式和单倍型结构、频率的地理变异以及CNVs独立起源的数量来做到这一点。最后，我们将进行一个CNV的重点调查-包含GTP环化水解酶位点-这可能是功能的重要性。我们将测试的假设，该基因的扩增补偿酶的代谢效率进一步下降的叶酸途径携带突变，赋予抗叶酸化合物的抗性。这将使用选择和转染实验结合药物抗性、适应性和通过叶酸途径的通量的测定来完成。这项研究与公共卫生的相关性。基因拷贝数的变化对寄生虫生物学有很大的影响。我们建议系统地描述疟原虫基因组中的DNA重排，并评估它们在这一重要病原体中所起的作用。

DESCRIPTION (provided by applicant): Gene copy number variation (CNV) is ubiquitous in eukaryotic genomes and the resultant dosage changes can play an important role in determining phenotype. We aim to define the extent, evolutionary dynamics and functionality of CNV in the malaria parasite genome (Plasmodium falciparum). To achieve this we will use comparative genomic hybridization (cGH) which provides an efficient tool for assaying both CNV and single nucleotide polymorphism (SNP) and is particularly effective in haploid genomes. Preliminary data reveal abundant CNV (both deletions and amplifications) in the P. falciparum genome and demonstrate accurate prediction of SNPs. The project will be divided into three parts. First, we will identify all CNV and catalogue flanking SNPs in a worldwide parasite collection. The populations sampled will comprise freshly isolated single-clone infections from Thailand, Papua New Guinea, Colombia and Gambia. We will measure the size, gene content, and arrangement and population frequency of genome rearrangements. Our second aim will be to evaluate evidence for adaptive evolution of genome regions showing CNV to highlight genome segments that could be influenced by recent evolutionary pressures such as selection by drugs. We will do this by examining patterns of linkage disequilibrium and haplotype structure in flanking SNP polymorphism, geographical variation in frequency, and by examination of numbers of independent origins of CNVs. Finally, we will conduct focused investigations of one CNV - containing the GTP cyclohydrolase locus - that is likely to be of functional importance. We will test the hypothesis that amplification of this gene compensates for reduced metabolic efficiency of enzymes further down the folate pathway carrying mutations that confer resistance to anti-folate compounds. This will be done using selection and transfection experiments in combination with assays of drug resistance, fitness, and flux through the folate pathway. Relevance of this research to public health. Variation in the number of gene copies can have a large influence on parasite biology. We propose to systematically describe DNA rearrangements in the malaria parasite genome, and evaluate the role that they play in this important pathogen.