利用全基因组外显子测序技术研究青少年特发性脊柱侧凸新的致病基因及相关发病机制

Adolescent idiopathic scoliosis (AIS) is a common spinal deformity that occurs in the puberty. In the past decades, a few common variants have been identified to be associated with the susceptibility of AIS by traditional candidate gene analysis and genome-wide association analysis. However, it is noteworthy that they can only explain less than 2% of the overall cause of AIS. For the complex diseases, whole-exome sequencing (WES) in the cohort of familial members and sporadic cases has become an efficient method to identify the rare variants related to the development of the disease. Meanwhile, it has been well documented that the rare variants can explain the causes of the disease more effectively as compared with the common variants. This project aims to detect mutations in a large cohort of AIS families using WES, which shall be further verified in more than 5000 sporadic AIS patients. Moreover, induced mutation in zebrafish model will be used to clarify the correlation between the mutation and the occurrence and progression of scoliosis. To summarize, these experiments will help discover new mutations involved in the development of AIS and further explore its pathogenesis.

青少年特发性脊柱侧凸（AIS）是一种好发于青春期的常见脊柱慢性疾病。尽管传统候选基因法和全基因组关联分析已定位十余个AIS易感基因多态性位点，它们仅能解释不到2%的疾病发生原因。利用家系结合散发病例进行全外显子组测序（WES）研究目前已成为人类复杂性疾病的热门研究手段。WES有助于高效准确地确定疾病发生相关的稀有突变，而稀有突变位点较易感多态性位点可更有效地解释疾病发生原因。本项目计划采用WES测序技术对数十个AIS家系进行基因突变的检测，并在逾5000例散发AIS患者中进一步验证。同时利用斑马鱼基因突变模型研究新发现的突变与脊柱侧凸发生及进展的相关性，上述研究将有助于进一步发现新的与AIS发生相关的基因突变并深入探索发病机制。

青少年特发性脊柱侧凸（AIS）是一种好发于青春期的常见脊柱遗传性疾病。尽管传统候选基因法和全基因组关联分析已定位十余个AIS发生易感多态性位点，它们仅能解释不到2%的疾病发生原因。本项目采用全外显子测序技术对数十个AIS家系合计96例患者进行基因突变的检测并在逾5000例散发AIS患者中进一步验证，有如下研究成果：..1.对于海量新发突变位点，我们首先采用了候选区域筛选法，即对已知的AIS易感区域进行重点筛查。在对SLC39A8外显子测序进行分析后，我们成功得到1个新的AIS致病编码位点rs11097773，位于SLC39A8基因的2号外显子，它可能会引起编码蛋白的错义突变。我们进一步检测了AIS患者椎旁组织中的SLC39A8表达，发现存在突变的患者该基因表达量显著下调。..2.斑马鱼脑脊液流速异常及纤毛发育受损是近年来AIS发病新的病因假说。本课题组在外显子测序基础上进一步分析了神经肽受体UTS2R基因外显子序列，发现数个新的突变。其中，我们成功得到2个新的AIS致病编码位点rs11654140和rs568196624，它们可能会引起编码蛋白的能量不稳定进而影响蛋白结构。我们进一步检测了AIS患者椎旁组织中的UTS2R表达，发现存在突变的患者该基因表达量显著下调。..3. 肌骨交叉作用在骨骼畸形发育中的作用是当前研究热点。AIS患者椎旁肌的发育及相关信号通路研究对解释该疾病成因有重要作用。本团队前期全基因组关联分析研究发现的易感基因LBX1在肌发育中起关键作用。通过整合本全外显子测序项目数据和先前全基因组关联分析数据，我们在LBX1启动子区域发现新的调控突变位点rs1322330，可通过影响转录因子EST-1的结合从而影响启动子转录活性。本团队进一步通过细胞学实验发现LBX1表达与患者II型肌纤维存在显著相关性。通过提纯和培养患者肌卫星细胞，本研究证实了过表达LBX1可影响肌细胞增殖和分化。.上述新发现均以原创性论著形式发表于SPINE杂志和Frontiers in Cell and Developmental Biology杂志。为AIS遗传病因学研究提供了新的思路，也为未来临床转化应用奠定相关理论基础。