Variations Of RIPPLY2 Gene In Congenital Scoliosis

Background:Congenital Scoliosis (CS) is a spinal deformity with a spinal coronary curvature of 10°or greater caused by abnormal vertebrate development during embryo. CS is characterized by severe deformity, rapid development and multiple complications. It is one of the most important causes of adolescent disability, carrying with high burden to the family and the society.Somitogenesis is a very conserved and important process in which somite is periodically formed from the presomitic mesoderm (PSM). Each somite then differentiates into vertebrae, paravertebral muscles and dermis. Abnormal somitogenesis is the reason of congenital vertebral malformation. However, the definite etiology of CS still remains to be elucidated. Without effective method for its early prediction CS is treated with braces and operation in order to prevent rapid progress of the disease. Therefore, it is of significant importance to find the etiology, establish methods of early diagnosis and intervention, and reduce the occurrence of this high pathogenic spinal deformity. Genetic etiologies for CS is gradually accepted. In 2015, our research team identified that Copy Number Variation (CNV), together with Single Nucleotide Variation (SNV), causes hemivertebrae, accounting for 11% of its etiology. This finding implies both CNV and SNV are important in genetic etiology studies.In 2005, ripply family was found to be of importance during somitogenesis. RIPPLY2 gene regulate TBX6 in a negative feedback loop, coordinating somitogenesis. In 2015, two consecutive clinical reports indicate that RIPPLY2 may be important causes of CS. However, its contribution to CS is still unknown.Objects:To investigate SNVs and CNVs of RIPPLY2 gene so as to elucidate the mechanism and validate its contributions to CS.MethodsScreening SNVs and CNVs of RIPPLY2 gene with Next Generation Sequencing technology among 109 CS patients with hemivertebrae. Further more, Whole Genome Sequencing for the trio of one patient with rare RIPPLY2 CNV was conducted. Raw data was annotated after strict quality control. Variants with low allele frequency and high predicted pathogenicity was analyzed.ResultsTotally 109 cases with hemivertebrae (including CS type Ⅰ and Ⅲ) were screened with next generation sequencing and 10 cases of TBX6 null mutation were found and excluded from this analysis. Among the remaining 99 cases,3 SNVs were identified, namely c.17G>A (p.G6D), c.29C＞A (p.T10K) and c.177G>A (p.M59I), all of which were heterozygous missense mutations. They were novel in public databases. One duplication contatining the RIPPLY2 gene was identified. Whole Genome Sequencing and PCR for the CNV trio confirmed the validity and its maternal origin of the duplication. No additional highly deleterious variant from her father which may regulate RIPPLY2 expression was identified.ConclusionsSNVs and CNVs of the RIPPLY2 gene are associated with CS with hemivertebrates, awaiting further investigation with bigger sample size.