Mechanism of subtelomeric breaks

描述(申请人提供)：拷贝数变异(CNV)是基因组多样性和人类疾病的一个重要来源9，10.亚端粒CNV是发生在染色体末端的大的缺失、重复和易位，导致3-6%的特发性智力低下病例6，11-13。有亚端粒重排的人有智力残疾、自闭症、畸形特征和/或其他出生缺陷。使用细胞遗传学测试，如亚端粒荧光原位杂交(FISH)或阵列比较基因组杂交(CGH)，儿童通常被诊断为亚端粒重排。据估计，在受影响的儿童中检测到的致病性CNV中，亚端粒重排的比例高达30%9，14，亚端粒重排的总体流行率估计为1/10,00015。尽管对人类健康有影响，但人们对引起亚端粒CNV的染色体断裂和修复机制知之甚少。研究新生血管形成的基础生物学对于了解致病结构重排的原因和相关的危险因素至关重要。我们假设特定的DNA序列很容易被破坏。为此，我们建议分离端粒下断裂位点下的人类DNA序列，并从功能上剖析导致双链断裂(DSB)的基序。这项建议的目的是1)精细定位亚端粒断裂点，2)识别导致基因组不稳定的亚端粒序列基序，以及3)在酵母总染色体重排(GCR)试验中对断裂基序进行功能注释，以定量亚端粒序列中的染色体断裂。综合基因组学、生物信息学和体内断裂实验将确定DNA序列是否在亚端粒断裂中发挥作用。我们的研究还将捕捉亚端粒重排的基因组结构，这将使我们能够确定作用于亚端粒DSB的修复机制。这一提议有望揭示促进基因组不稳定的新序列基序和DNA修复的新机制。这些数据对于理解在整个人类基因组中形成亚端粒重排和其他CNV的力量是至关重要的。 与公共卫生相关：发生在染色体末端的亚端粒重排是导致智力残疾和出生缺陷的重要原因。我们建议对亚端粒断裂点进行测序，并确定促进基因组不稳定性的序列基序。我们将通过将亚端粒序列基序引入测量染色体断裂的酵母试验来确定它们的脆弱性。

DESCRIPTION (provided by applicant): Copy number variation (CNV) is a significant source of genomic diversity and human disease9,10. Subtelomeric CNVs are large deletions, duplications, and translocations that occur at chromosome ends and are responsible for 3-6% of idiopathic mental retardation cases6,11-13. Individuals with subtelomeric rearrangements have intellectual disabilities, autism, dysmorphic features, and/or other birth defects. Children are typically diagnosed with a subtelomeric rearrangement using cytogenetic tests such as subtelomeric fluorescence in situ hybridization (FISH) or array comparative genomic hybridization (CGH). Subtelomeric rearrangements have been estimated to account for up to 30% of pathogenic CNVs detected in affected children9,14, and the overall prevalence of subtelomeric rearrangements is estimated at 1/10,00015. Despite the impact on human health, very little is known about the mechanisms of chromosome breakage and repair that give rise to subtelomeric CNV. Research on the fundamental biology of CNV formation is critical to understanding the causes of and risk factors associated with disease-causing structural rearrangements. We hypothesize that particular DNA sequences are susceptible to breakage. To this end, we propose to isolate human DNA sequences that underlie subtelomeric breakage sites and functionally dissect the motifs that cause double-strand breaks (DSBs). The aims of this proposal are to 1) fine-map subtelomeric breakpoints, 2) identify subtelomeric sequence motifs that contribute to genomic instability, and 3) functionally annotate breakage motifs in a yeast gross chromosomal rearrangement (GCR) assay that quantifies chromosome breakage in the subtelomeric sequence. Integrated genomic, bioinformatics, and in vivo breakage experiments will determine whether or not DNA sequence plays a role in subtelomeric breakage. Our studies will also capture the genomic structure of subtelomeric rearrangements, which will allow us to determine the mechanisms of repair acting on subtelomeric DSBs. This proposal promises to reveal new sequence motifs that promote genomic instability and novel mechanisms of DNA repair. These data will be critical to understanding the forces that shape subtelomeric rearrangements and other CNV throughout the human genome. PUBLIC HEALTH RELEVANCE: Subtelomeric rearrangements, which occur at the ends of chromosomes, are a significant cause of intellectual disability and birth defects. We propose to sequence subtelomeric breakpoints and identify sequence motifs that promote genomic instability. We will determine the fragility of subtelomeric sequence motifs by introducing them into a yeast assay that measures chromosome breakage.