Genetic mechanism of conserved ancestral haplotype in SCA10

项目摘要/摘要 脊髓小脑型共济失调10(SCA10)是一种罕见的常染色体显性遗传性共济失调，由 ATXN10基因内含子9的(ATTCT)n重复。SCA10仅存在于拉丁美洲(洛杉矶) 美洲原住民血统和东亚(EA)人口。我们所有的SCA10患者都有一个 包括rs41524745上的G等位基因的单一单倍型。G等位基因的次要等位基因频率为2- 在EA和LA人群中为4%，而在其他地方为0%。这些数据表明，单一的祖先突变具有 在美洲出现之前，在15,000-20,000年前在EA兴起于G(+)单倍型。尽管 Rs41524547和sca10重复序列之间的距离足够长，可以进行多次历史重组 在正常人群中发生的事件，我们的SCA10人群没有显示G等位基因与SCA10的分离 扩张。此外，G等位基因位于miR4762-5p的种子序列附近。我们获得了 从1000个基因组库和我们自己的普通人群样本中提取G(+)DNA，并进行检测 SCA10扩展。我们发现，在这些样本中，多达25%的样本显示出SCA10重复扩展。我们的研究 进一步表明，被(ATTCC)n或(ATCCT)n(ATCCC)n重复中断的SCA10扩展是完全的 穿透性纯(ATTCT)n膨胀显示渗透率降低。基于这些观察，我们 建议检验以下假设：(1)rs41524547上的G等位基因使SCA10重复为 纯(ATTCT)n重复膨胀(A类膨胀)，基本上保持非渗透性，以及(2) (ATTCT)n-(ATTCC)n(类型B)或(ATTCT)n-(ATCCT)n-(ATCCC)n(类型C)重复插入A型 扩张驱动了SCA10的致病性。为了检验这些假设，我们将致力于： 目的1)确定SCA10扩增与rs41524547的G等位基因的关系： (1)病例对照研究，证明SCA10扩增与G(+)单倍型密切相关。 (1B)确定SCA10患者和其他G(+)个体的重复结构。 目的2)证实B型和C型扩张是完全致病的，而不是A型扩张。 (2a)鉴定分别表达A、B和C型扩展的转基因小鼠的表型。 (2B)确定SCA10患者iPS细胞来源的浦肯野样细胞(PC)的表型 A、B和C展开。 (2C)在50岁的≥中建立A、B和C型扩展的外显率。 (2D)获得具有A、B和C型扩展的SCA10家系成员的横断面临床数据。 目的3)确定rs41524547的G等位基因是否降低下游重组率，保护 对抗SCA10 RNA扩展的毒性，或促进SCA10重复序列的扩展状态： (3)测定SCA10患者精子rs41524547-SCA10区间的重组率。 (3B)确定rs41524547处G-to-C基因组编辑在IPSC来源的SCA10 PC中的效果。

PROJECT SUMMARY/ABSTRACT Spinocerebellar ataxia type 10 (SCA10) is a rare autosomal dominant ataxia caused by a large expansion of (ATTCT)n repeat in intron 9 of the ATXN10 gene. SCA10 exists exclusively in Latin American (LA) populations of Native American Ancestry and East Asian (EA) populations. All our SCA10 patients share a single haplotype that includes the G allele at rs41524745. The G allele has the minor allele frequency of 2- 4% in EA and LA populations but 0% elsewhere. These data suggest that the single ancestral mutation has risen on the G(+) haplotype in EA 15,000-20,000 years ago before peopling Americas. Although the distance between rs41524547 and the SCA10 repeat is long enough for multiple historical recombination events in normal populations, our SCA10 population shows no dissociation of the G allele from the SCA10 expansion. Furthermore, the G allele is located adjacent to the seed sequence of miR4762-5p. We obtained G(+) DNA from the 1000 Genomes repository and our own general population samples and tested for SCA10 expansions. We found up to 25% of these samples show SCA10 repeat expansions. Our studies further suggest that SCA10 expansions interrupted by (ATTCC)n or (ATCCT)n(ATCCC)n repeat are fully penetrant while pure (ATTCT)n expansions show reduced penetrance. Based on these observations we propose to test following hypotheses: (1) the G allele at rs41524547 predisposes the SCA10 repeat for pure (ATTCT)n repeat expansion (Type A expansion), that remains mostly non-penetrant, and (2) the (ATTCT)n-(ATTCC)n (Type B) or (ATTCT)n-(ATCCT)n-(ATCCC)n (Type C) repeat insertion into Type A expansion drives the SCA10 pathogenicity. To test these hypotheses we will aim to: Aim 1) Determine the relationship between SCA10 expansions and the G allele at rs41524547: (1a) Case-control study to document the tight association of SCA10 expansion with the G(+) haplotype. (1b) Determine repeat structures in SCA10 patients and other G(+) individuals. Aim 2) Confirm that Type B and Type C, but not Type A, expansions are fully pathogenic. (2a) Characterize phenotypes of transgenic mice expressing each of Types A, B and C expansion. (2b) Determine phenotype of Purkinje-like cells (PC) derived from iPS cells of SCA10 patients with Types A, B and C expansions. (2c) Establish penetrance rates of Type A, B and C expansions in SCA10 sibships ≥50 years of age. (2d) Obtain cross-sectional clinical data from SCA10 family members with Types A, B and C expansions. Aim 3) Determine if the G allele at rs41524547 reduces downstream recombination rates, protects against the toxicity of SCA10 RNA expansions, or promote expanded states of the SCA10 repeat: (3a) Determine the recombination rate in the rs41524547-SCA10 interval in sperm from SCA10 patients. (3b) Determine the effect of G-to-C genome editing at rs41524547 in iPSC-derived SCA10 PC.