The mechanistic basis for constitutional MLH1 methylation (epimutation)

项目摘要 林奇综合征是最常见的遗传性癌症，通常由可遗传基因引起。 影响DNA错配修复基因之一的突变，最常见的是位于3号染色体上的MLH1。 所谓的“MLH1突变”是LS的另一种原因。MLH1表型突变的特征是 在整个正常过程中，MLH1启动子的单拷贝甲基化伴随着基因活性丧失 纸巾。这使携带者更容易患上LS型癌症。患者的身份由 检测血液或任何其他正常组织DNA中的MLH1甲基化。对中国传统文化的研究 患有MLH1突变的病例的家庭成员表现出明显的遗传模式，因此 不同的风险，从父母传给后代，如果有的话。因此，MLH1表型突变 在人类中提供了一个独特的机会来研究表观遗传变化是如何发生并从而导致 疾病，以及表观遗传变化是如何在一些家庭中从一代传到下一代的， 但在其他人身上却并非如此。希钦斯博士是这项研究的首席研究员，他收集了一组独特的 MLH1突变是癌症的原因，并建立了该基因的遗传模式。 在过去的13年里，她通过在这一领域的工作，在每个家庭中都得到了赞誉。这项研究的目标是 通过对基因组的深入研究来确定MLH1半突变的机制 和患者及其家属的表观基因组。在目标1中，我们将使用最先进的链接-阅读全文 基因组测序“对选定的患者和家庭成员的整个基因组进行排序，以确定 3号染色体或其他染色体上的各种类型的基因突变是MLH1发病的基础 上变种。在目标2中，我们将确定MLH1是否是唯一受到错误宪法影响的基因 甲基化，或者如果其他基因同时甲基化。这将为我们提供关于 表观突变是由于广义表观遗传扰动或影响MLH1的焦点机制引起的 独自一人。在目标3中，我们将验证任何已确定的遗传或表观遗传改变，并确定它们的分离 家族中的模式。我们还将对之前和2003年确定的遗传变异进行功能分析 目标1.总的来说，这些研究将为已确定的基因改变之间的因果联系提供证据 和MLH1突变，使我们能够预测世代之间遗传的可能性。最后， 我们将研究患者白细胞被诱导成多能状态，以模拟表观遗传重新编程。 这些患者在MLH1发生的事件。这将阐明为什么一些表位突变是遗传的 其他人则不是。如果成功，这项研究将使我们能够改进临床管理和遗传咨询 有易患癌症的MLH1基因突变的家庭。更广泛地说，这项研究将提供重要的新的 对致病遗传状态和表观遗传状态之间的相互作用以及如何改变的见解 表观遗传状态可能会从一代传到下一代，或者在几代之间被抹去。

Project Summary Lynch syndrome (LS), the most common hereditary cancer condition, is usually caused by a heritable genetic mutation affecting one of the DNA mismatch repair genes, most frequently MLH1, located on chromosome 3. So-called “MLH1 epimutation” is an alternative cause for LS. MLH1 epimutations are characterized by methylation of a single copy of the MLH1 promoter, accompanied by loss of gene activity, throughout normal tissues. This predisposes carriers to the development of LS-type cancers. Patients are identified by the detection of “constitutional” MLH1 methylation in DNA from blood or any other normal tissue. Studies of the family members of cases with an MLH1 epimutation have revealed distinct patterns of inheritance and hence differing risks of passing the epimutation from parent to offspring, if at all. Therefore, MLH1 epimutations provide a unique opportunity in humans for studying how epigenetic alterations arise and thereby cause disease, and also how epigenetic alterations are transmitted from one generation to the next in some families, but not in others. Dr. Hitchins, the principle investigator of this study, has assembled a unique collection of families with an MLH1 epimutation as the cause for cancer, and has established the inheritance patterns of the epimutation in each family, through her work in this field over the past 13 years. The goals of this study are to determine the mechanisms that underlie MLH1 epimutation by undertaking an in-depth study of the genomes and epigenomes of the patients and their families. In Aim 1, we will use state of the art “linked-read whole genome sequencing” to sequence the entire genome of selected patients and family members to determine if various types of genetic mutations on chromosome 3, or other chromosomes, underlie the onset on MLH1 epimutations. In Aim 2, we will determine if MLH1 is the only gene that is subject to erroneous constitutional methylation, or if other genes are simultaneously methylated. This would provide insights into whether epimutations arise because of a generalized epigenetic perturbation, or a focal mechanism affecting MLH1 alone. In Aim 3, we will validate any genetic or epigenetic alterations identified and determine their segregation patterns in families. We will also perform functional analyses of genetic variants identified previously and in Aim 1. Collectively, these studies will provide evidence for a causal link between genetic alterations identified and the MLH1 epimutation, and enable us to predict the likelihood of inheritance between generations. Finally, we will study patient white blood cells induced into a pluripotent state to model the epigenetic reprogramming events that occur at MLH1 in these patients. This will shed light on why some epimutations are inherited and others not. If successful, this study will enable us to improve the clinical management and genetic counselling of families with a cancer-predisposing MLH1 epimutation. More broadly, this study will provide significant new insights into the interaction between disease-causing genetic and epigenetic states, as well how altered epigenetic states may are passed from one generation to the next, or erased between generations.