FLNC和PDE4D基因突变在一例新生儿先天性多发畸形中的机制研究

随着高通量测序技术的发展，全外显子组测序技术（WES）已成为临床遗传病诊断的有效工具。本课题组前期通过WES发现一例先天性多发畸形患儿同时携带FLNC基因P2217H和PDE4D基因I277V两个杂合新发突变。虽然FLNC与PDE4D基因在胚胎发育中占重要地位，但这两个基因的2个突变均未报道与该患儿的临床表型相关。因此，本课题拟：分别构建FLNC和PDE4D突变体和双重突变体的斑马鱼模型，观察这两个基因突变在斑马鱼胚胎对称发育中的角色和作用；并在体外采用人ES-H9细胞株，转染FLNC、PDE4D突变体和双重突变体，向中胚层诱导分化，观察内脏异位相关因子NODAL、LEFTY2等的表达和分布情况。本课题可阐明FLNC和PDE4D基因突变在胚胎不对称发育中的作用，明确FLNC和PDE4D基因是否为内脏异位的候选基因，并为解释本例患儿的临床表型提供理论依据，为早期预防及治疗、遗传咨询提供线索。

With the development of high-throughput sequencing technology, the whole exome sequencing (WES) has been widely used in the clinical diagnosis of genetic diseases. Two de novo heterozygous mutations of FLNC and PDE4D (FLNC: c.C6650A, p.P2217H; PDE4D: c.A829G, p.I277V) were found in a neonate with multiple congenital anomaly by using WES. Although FLNC and PDE4D gene play important roles in embryonic development, the two mutations of these two genes were not reported be associated with the clinical phenotype of the children. Therefore, we design this project to do a study: Firstly, the FLNC and PDE4D mutant and double mutant will be constructed in zebrafish model. The purpose is to observe the role and function of the two genetic mutations in symmetric zebrafish embryo development. Secondly, in vitro, FLNC, PDE4D mutants and double mutant will be established in human ES-H9 cell lines. Differentiation experiment will be performed to induced to mesoderm. The purpose is to observe the expression and distribution of heterotaxia related key factor, NODAL、LEFTY2 and PITX2C. Through this study, we will clarify the role of FLNC and PDE4D mutant in embryonic asymmetric development, and clear whether PDE4D and FLNC gene are a candidate gene for heterotaxy. So, we can provide evidence to explain the cause of the clinical phenotype in this case, and can find clues for the genetic counseling, prenatal screening and early preventive treatment.