The Phenotype And Pathogenic Mechanisms Studies Of The TBX6 Associated 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 charatered by rapid development, severe deformity and multiple complications. It is one of the most causes of adolescent disability and brings high burden to the family and the society.The spine of the vertebrate is developed from the presomitc mesoderm (PSM) from which somite is periodically developed, this process is called somitogenesis. Abnormal somitogenesis is the reason of congenital vertebrate deformity. However, the definite etiology of CS is obscure and there is still no effective method for its early prediction. Clinially, the treatment of CS is mainly focused on bracement and operation in order to avoid the rapid progress of the disease. Therefore, it is important to find the etiology and to reduce the occurrence of this high pathogenic spinal deformity.The abnormal of genes and changes of environmental factors in embryo or during labor are the main casue of CS. With more and more genetic etiology were reported, the genetic influence of CS is gradually remarked. CS is a complicated disease with multiple genes and multi genetic models related. Little larege sample of genetics research of CS was reported because it is difficult to collect large scale of patients with the same phenotype as patients are usually infertile, furthermore, the strategy of analysis based on the mouse-human synteny restricts the profound and creative study as the real mechanism of molecular biological etiology of CS is unknow either.CNV (copy number variation), which is defined as deletions or duplications in genome DNA, can lead to human diseases including congenital abnormalities. CNVs can be classified into recurrent CNVs and non-recurrent CNVs as the sizes and boundaries of recurrent CNVs were relatively constant. Many CNVs are inherited polymorphisms and account for a significant proportion of the healthy human genome.There is an interplay between CNV detection and array platform resolution. A genomewide analysis of copy-number variants was performed in 20 trios (consisting of a person with sporadic congenital scoliosis and two healthy parents) with the use of Agilent 1 × 1M CGH microarrays. We identified recurrent deletions in proximal 16p11.2 in two persons. Both of these heterozygous deletions were de novo.Objects:To investigate the genetic etiology, clinical phenotype and pathogenic mechanisms of TBX6-associated CS, and to establish the animal modelMethodsWe conducted genetic analyses in 237 sporadic CS patients, and 166 Chinese controls using comparative genomic hybridization, quantitative PCR, and DNA sequencing. Phenotypes were compared between the TBX6-associated CS and non-TBX6-associated CS. Zebrafish model of TBX6-associated CS was builded by CRISPR-Cas9 to verify the phenotypic characteristics and pathogenic mechanisms.Results23 heterozygous TBX6 null mutations were identified in 237 sporadic CS, including copy-number variants (17 16p11.2/TBX6 deletions) and single-nucleotide variants (one nonsense and five frameshift mutations). However, the discordant intra-familiar phenotypes observed in two 16p11.2/TBX6 deletion pedigrees suggest that heterozygous null TBX6 is insufficient to cause CS. Instead, we identified a common TBX6 haplotype as the second risk allele, which was shared by all 23 carriers of TBX6 null mutations. Notably, hemivertebra, simple rib malformation and less NTD involved are the common clinical subtype of TBX6-associated CS. The zebrafish model showed a gradient of vertebral deformities, depending on the degree of mutation of tbx6 and tbx16.ConclusionsThe TBX6 model accounts for 10% of CS in the studied populations, representing a significant CS locus. We defined CS with 16p 11.2 deletion/TBX6 nonsense mutations with T-C-A hypomorphic as TBX6-associated congenital scoliosis, which has its unique characteristics in clinical phenotype. In the zebrafish model, pathogenesis and phenotype correlations are in line TBX6 CS dose model, but the exact mechanism remains to be further studied.