Sanchez-Flores Marina, Corral-Juan Marc, Gasch-Navalón Esther, Cirillo Davide, Sanchez Ivelisse, Matilla-Dueñas Antoni
Neurogenetics Unit, Department of Neuroscience, Germans Trias i Pujol Research Institute (IGTP), Universitat Autònoma de Barcelona-Can Ruti Campus, Carretera de Can Ruti, Camí de les Escoles s/n, 08916, Badalona, Spain.
Barcelona Supercomputing Center (BSC), Barcelona, Spain.
Hum Genet. 2024 Mar;143(3):211-232. doi: 10.1007/s00439-024-02644-7. Epub 2024 Feb 23.
Spinocerebellar ataxia subtype 37 (SCA37) is a rare disease originally identified in ataxia patients from the Iberian Peninsula with a pure cerebellar syndrome. SCA37 patients carry a pathogenic intronic (ATTTC)n repeat insertion flanked by two polymorphic (ATTTT)n repeats in the Disabled-1 (DAB1) gene leading to cerebellar dysregulation. Herein, we determine the precise configuration of the pathogenic 5'(ATTTT)n-(ATTTC)n-3'(ATTTT)n SCA37 alleles by CRISPR-Cas9 and long-read nanopore sequencing, reveal their epigenomic signatures in SCA37 lymphocytes, fibroblasts, and cerebellar samples, and establish new molecular and clinical correlations. The 5'(ATTTT)n-(ATTTC)n-3'(ATTTT)n pathogenic allele configurations revealed repeat instability and differential methylation signatures. Disease age of onset negatively correlated with the (ATTTC)n, and positively correlated with the 3'(ATTTT)n. Geographic origin and gender significantly correlated with age of onset. Furthermore, significant predictive regression models were obtained by machine learning for age of onset and disease evolution by considering gender, the (ATTTC)n, the 3'(ATTTT)n, and seven CpG positions differentially methylated in SCA37 cerebellum. A common 964-kb genomic region spanning the (ATTTC)n insertion was identified in all SCA37 patients analysed from Portugal and Spain, evidencing a common origin of the SCA37 mutation in the Iberian Peninsula originating 859 years ago (95% CI 647-1378). In conclusion, we demonstrate an accurate determination of the size and configuration of the regulatory 5'(ATTTT)n-(ATTTC)n-3'(ATTTT)n repeat tract, avoiding PCR bias amplification using CRISPR/Cas9-enrichment and nanopore long-read sequencing, resulting relevant for accurate genetic diagnosis of SCA37. Moreover, we determine novel significant genotype-phenotype correlations in SCA37 and identify differential cerebellar allele-specific methylation signatures that may underlie DAB1 pathogenic dysregulation.
脊髓小脑共济失调37型（SCA37）是一种罕见疾病，最初在伊比利亚半岛患有纯小脑综合征的共济失调患者中被发现。SCA37患者在失活-1（DAB1）基因中携带一个致病性内含子（ATTTC）n重复插入序列，两侧为两个多态性（ATTTT）n重复序列，导致小脑调节异常。在此，我们通过CRISPR-Cas9和长读长纳米孔测序确定了致病性5'（ATTTT）n-（ATTTC）n-3'（ATTTT）n SCA37等位基因的精确结构，揭示了它们在SCA37淋巴细胞、成纤维细胞和小脑样本中的表观基因组特征，并建立了新的分子和临床相关性。5'（ATTTT）n-（ATTTC）n-3'（ATTTT）n致病性等位基因结构显示出重复序列不稳定性和差异甲基化特征。疾病发病年龄与（ATTTC）n呈负相关，与3'（ATTTT）n呈正相关。地理起源和性别与发病年龄显著相关。此外，通过机器学习，考虑性别、（ATTTC）n、3'（ATTTT）n以及SCA37小脑样本中7个差异甲基化的CpG位点，获得了关于发病年龄和疾病进展的显著预测回归模型。在分析的所有来自葡萄牙和西班牙的SCA37患者中，鉴定出一个跨越（ATTTC）n插入序列的964kb常见基因组区域，证明了伊比利亚半岛SCA37突变的共同起源，该起源可追溯到859年前（95%置信区间647-1378）。总之，我们展示了对调控性5'（ATTTT）n-（ATTTC）n-3'（ATTTT）n重复序列长度和结构的准确测定，通过CRISPR/Cas9富集和纳米孔长读长测序避免了PCR偏倚扩增，这对于SCA37的准确基因诊断具有重要意义。此外，我们确定了SCA37中新的显著基因型-表型相关性，并鉴定出可能是DAB1致病性调节异常基础的小脑等位基因特异性差异甲基化特征。
