Functional characterization of non-coding variants in disease-causing regions of benign choreoathetosis (CAHTP) and thyroid dysgenesis (CHTD)

转录因子NKX2-1不仅是甲状腺滤泡细胞的主要调节因子，而且在腹侧端脑、间脑和肺泡上皮细胞中也具有特殊的神经细胞类型。NKX2-1基因突变会导致良性舞蹈手足徐动症和先天性甲状腺功能减退伴或不伴肺功能障碍(CAHTP，OMIM#610978)，这是一种常染色体显性遗传性疾病，具有不同的外显率和甲状腺和肺表型的严重程度；正如我们小组在2002年首次展示的那样。在第一个资助期间，我们使用WGS来识别NKX2-1基因完整的CAHTP患者队列中NKX2-1基因位点的非编码变异。通过分析NKX2-1基因座的表观基因组特征，我们在NKX2-1基因下游的14q13处指定了一个大约100kb的疾病相关区域。在这个项目的第一部分，我们将使用不同的技术来定位这个区域，以检查这个基因组区域的顺式调节作用，并分析WGS在我们的25名CAHTP索引患者队列中检测到的非编码变异的影响。为了更深入地了解甲状腺发育不良(CHTD)的遗传机制，我们与日本东京庆应义乌大学医学院的Satoshi Narumi进行了一项合作的全基因组关联研究(Gwas)，并在2q33.3的非编码基因组区域确定了CHTD的第一个疾病相关区域。重要的顶部基因SNP(Rs9789446)位于72kb的内含子区域，该区域包含FZD5和CCNYL1在甲状腺组织中表达的顺式调控序列，根据表观基因组、转录组和染色质相互作用数据集的组合推断。为了正式证明这个非编码区与甲状腺发育不全的疾病相关性，我们将在本项目的第二个资助期中鉴定2q33.3处的顺式调控元件的活性。

The transcription factor NKX2-1 is a master regulator not only of thyroid follicular cells, but also of specific neuronal cell types in the ventral telencephalon, the diencephalon, and of pulmonary alveolar epithelial cells. Mutations of the NKX2-1 gene cause benign choreoathetosis and congenital hypothyroidism with or without pulmonary dysfunction (CAHTP, OMIM #610978), an autosomal dominant disorder with variable penetrance and severity of thyroid and pulmonary phenotypes; as initially shown by our group already in 2002. During the first funding period we used WGS to identify non-coding variants in the NKX2-1 gene locus of a cohort of CAHTP patients with an intact NKX2-1 gene. By analyzing epigenomic signatures of the NKX2-1 gene locus, we assigned a putative 100 kb disease-associated region at 14q13 downstream of the NKX2-1 gene. In the first part of this project, we will target this region with different techniques to examine the cis-regulatory role of this genomic region and to analyze the impact of the non-coding variants detected by WGS in our cohort of 25 CAHTP index patients. To gain additional insights into the genetic mechanisms of thyroid dysgenesis (CHTD), we conducted a collaborative genome-wide association study (GWAS) with Satoshi Narumi from Keio University School of Medicine, Tokyo (Japan) and identified a first disease-associated region for CHTD in a non-coding genomic region at 2q33.3. The significant top genotyped SNP (rs9789446) was located in a 72 kb intronic region that contains cis-regulatory sequences for FZD5 and CCNYL1 expression in thyroid tissue as inferred from a combination of epigenome, transcriptome, and chromatin interaction data sets. To begin to formally demonstrate the disease relevance of this non-coding region for thyroid dysgenesis, we will characterize the activity of putative of cis-regulatory elements at 2q33.3 in the second funding period of this project.