The Dynamics of Mitochondrial Mutations

 描述（申请人提供）：mtDNA突变导致>200种遗传疾病。八分之一的无关女性是mtDNA疾病的携带者。大多数mtDNA疾病是异质性的-对他们来说，疾病相关的等位基因与野生型等位基因一起存在。mtDNA疾病的严重程度取决于组织中疾病相关等位基因的频率。因此，研究mtDNA等位基因频率在世代之间和人体内的变化至关重要。这些变化是由突变、遗传漂变和选择所控制的。然而，这些基本参数和相对贡献 形成mtDNA遗传组成的进化过程既没有被建模，也没有被详细描述。本研究的目的是探讨个体mtDNA等位基因频率在世代之间和组织之间变化的进化过程。为了实现这一目标，我们创建了一个独特的资源-从人类人群（来自宾夕法尼亚州中部）的母亲和两个孩子（母亲-两个孩子）三人组的口腔和血细胞中收集样本。我们将使用新开发的计算和统计工具对他们的mtDNA进行测序和分析，这些工具打包在一个可复制的软件管道中，将与科学界共享。在目标1中，我们将开发一种新的mtDNA突变和漂移的群体遗传学框架，并将其应用于200个母亲-2个孩子三人组的mtDNA序列。我们将估计生殖系，胚胎和体细胞突变率和瓶颈大小。需要估计生殖系瓶颈大小来预测儿童中的异质性水平，并且需要估计胚胎瓶颈大小来解析异质性在组织中的分布。生殖系突变率估计将有助于人类进化研究，体细胞突变率估计将为我们提供有关mtDNA疾病中突变累积的信息。目的2将测试年龄对女性生殖系中mtDNA突变积累的潜在影响。除了母子三人组之外，我们还将直接检查女性生殖系-通过对100名不同年龄女性未受精卵母细胞的mtDNA进行测序。mtDNA生殖系突变率可能会增加，因为卵母细胞老化过程会产生诱变环境。如果发现与年龄相关的mtDNA突变率增加，那么在现代西方社会中制定计划生育建议将非常重要，因为在现代西方社会中，生育经常被推迟。在目标3中，我们将开发新的基于似然的方法来检测mtDNA的选择。将这些方法应用于来自母子三人组和未受精卵母细胞的mtDNA测序数据，我们将比较生殖系与体细胞mtDNA选择的强度，并评估线粒体选择是否主要在细胞（或组织内的细胞）中的单个线粒体水平上或群体中的个体中起作用。这将大大有助于正在进行的辩论， 一个生物体的线粒体DNA选择开始起作用。因此，使用创新的方法，我们将解决有关mtDNA进化和疾病的关键问题。

 DESCRIPTION (provided by applicant): Mutations in mtDNA cause >200 genetic diseases. One in eight unrelated females is a carrier of an mtDNA disease. Most of mtDNA diseases are heteroplasmic - for them, the disease‐associated alleles are present alongside the wild‐type alleles. The severity of the mtDNA diseases depends on the frequency of the disease associated alleles in a tissue. Therefore, it is critical to study how mtDNA allele frequencies change between generations and within a human body. Such changes are governed by mutation, genetic drift, and selection. Yet the basic parameters and relative contribution of these evolutionary processes to shaping the mtDNA genetic makeup have neither been modeled nor characterized in detail. The goal of this proposal is to investigate evolutionary processes governing mtDNA allele frequency changes between generations and among tissues of an individual. To achieve this goal, we have created a unique resource - collected samples from buccal and blood cells of mother and two child (mother‐2child) trios from a human population (from Central Pennsylvania). We will sequence and analyze their mtDNA with newly developed computational and statistical tools packaged in a reproducible software pipeline that will be shared with the scientific community. In Aim 1 we will develop a novel population genetics framework for mtDNA mutation and drift, and will apply it to mtDNA sequences from 200 mother‐2 child trios. We will estimate germ‐line, embryonic, and somatic mutation rates and bottleneck sizes. The germ‐line bottleneck size estimate is needed to predict heteroplasmy levels in children, and the embryonic one to parse the distribution of heteroplasmies among tissues. The germ‐line mutation rate estimate will be useful for human evolutionary studies, and the somatic mutation rate estimate will inform us about mutation accumulation in mtDNA diseases. Aim 2 will test a potential effect of age on accumulation of mtDNA mutations in the female germ line. In addition to mother‐2 child trios, here we will examine female germ line directly - by sequencing mtDNA from unfertilized oocytes from 100 women of different ages. mtDNA germ‐line mutation rates might increase because of oocyte aging processes which create a mutagenic environment. An age‐related increase in mtDNA mutation rate, if found, will be important for formulating family planning recommendations in modern Western societies in which reproduction is frequently delayed. In Aim 3 we will develop novel likelihood‐based methods for detecting selection at mtDNA. Applying these methods to the mtDNA sequencing data from mother‐2child trios and unfertilized oocytes, we will contrast the strength of germ‐line vs. somatic mtDNA selection, and evaluate whether mitochondrial selection operates predominantly at the level of individual mitochondria in a cell (or cells within a tissue), or amon individuals in a population. This will significantly contribute to an ongoing debate about where in an organism mtDNA selection operates. Thus, using innovative methodology, we will address pivotal questions about mtDNA evolution and disease.