Neural Tube Defects Three Genetic Mutation Screening Phosphorylation Research And Three Vangl Gene Mutations

Neural tube defects(NTDs) occurred in very early stage during embryogenesis, caused by the failure of the neural tube to close properly that results in a group of syndromes, including anencephaly and myelomeningocele, craniorachischisis and spina bifida. Infants with anencephaly are still born or die shortly after birth, whereas many infants with spina bifida can survive. In addition to the emotional cost of spina bifida, the estimated monetary cost is also staggering. NTDs occurring in approximately one in1,000live births in the world wide, but occurred3%o live births in China, in several areas even as high as up to8%o, which constitute the second most frequent cause of human congenital abnormalities in China.Complex multigenetic causes and environmental causes have been suggested to contribute to NTDs. Recently, more and more research in mouse models of neural-tube defects indicated that nearly all of the genes involved in planar cell polarity(PCP) pathway play an important role in neural tube closure. The so-called PCP pathway core genes included:Frizzled(Fz), Dishevelled(Dsh/Dvl), Strabismus/Van Gogh(Stam/Vangl), Flamingo(Fmi), Prickle(Pk) and Diego(Dgo). In this study, we use case-control study methods to screen CELSR1and FUZZY, WNT11ease special rare mutations in163still born or miscarried Han Chinese fetuses with neural tube defects, aimed to investigate whether these mutations are associated with human neural tube defects through biochemical, genetics and model animal study. In addition, the present work was based on previous studys of three unique mutations of Vangl related with neural tube defects, which were found in the human neural tube defects cases.In163NTDs cases, we found ten missense mutations of the CELSR1gene: A1408T, N1884S, VI897I, C1898R, G1973D, L1995R, I2107V, P2385L, R2901H and E2903Q. By SNapShot genotyping methods, we identified five unique mutations: A1408T, N1884S, VI897I, G1973D and R2901H. While by the analysis of the conserved vertebrate amino acid sequence of CELSR1gene, it reveals that the five missense mutations are not very conservative. These mutations may be associated with human neural tube defects.But in163cases of neural tube defects, we found a rare mutation of WNT11and FUZZY genes. We found three mutations in WNT11gene:C.374G>A, C.408C>T, C.1037G>T. In addition, C.408C>T and C.1037G>T were synonymous mutations. C.374G>A was missense mutation. We found one synonymous mutation of FUZZY gene:C.1199C>T. It can be detected in13case samples. Because of the quantity limitation of the neural tube defect cases, we did not identify some rare mutations associated with the neural tube defects.As to three mutations:G39S, N313S, S84F of VANGL, we had successfully constructed the corresponding expression vectors carrying the three mutation sites, where mutant VANGL were fused Myc(in pCMV6-Myc vector) and GFP(Green fluorescent(in pEGFP-N2. Cell transfection and western blot experiments revealed that these three point mutations didn’t up-regulated the phosphorylation level of the encoded proteins, but the proteins expression were enhanced.Conclusion:Our findings, together with other early reports, suggest that rare mutations of the PCP-related genes may make a great contribution to human NTDs. It may have important clinical significance to the further elucidation of the genetic mechanism of NTDs.