Single Nucleotide Polymorphisms In5,10-methylenetetrahy-Drofolate Reductase(MTHFR)Gene And Preterm Birth

[Abstract] Preterm birth (PTB) is defined as any birth before37weeks of gestation, or less than259days of pregnancy, beginning the first day of a last menstrual period. The incidence of PTB in our country is about8%, and the trend is ascendant. PTB is the leading cause of perinatal morbidity and mortality. Currently, the cause of preterm birth is not entirely clear. It is generally agreed that PTB is caused by genetic factors and environmental factors. Epidemiological survey indicate that folic acid supplements before and during pregnancy can reduce the incidence of PTB. Therefore, lack of folic acid utilization ability has become an important direction in PTB genetic factors. The single nucleotide polymorphisms (SNPs) of5,10methylenetetrahydrofolate reductase (MTHFR) which is an important key enzyme in folate metabolisms become a hot focus. However, there are no studies on3’-untranslational region (3’-UTR) of MTHFR playing important rloe in gene regulation while functional SNPs searching is focused more on its coding regions. In addition, the results of association studies on PTB and coding regions are inconsistent. To look for genetic causes of PTB, this study focuses on3’-UTR which can regulate gene dose and exploring the association between PTB and SNPs in3’-UTR.The association between PTB and SNPs in3’-UTR of MTHFR gene has not been reported. We predicted microRNA (miRNA) interacted with3’-UTR of MTHFR by using online software (TargetScan, DIANA-micro and PicTar) and selected intersections of these three online softwares as candidate miRNAs. Then we used miRNASNP to predict SNP sites in the seed sequence of miRNA. Finally we got9 SNPs as candidate SNP sites. We used SNaPshot genotyping for the9candidate SNPs in480preterm samples and655normal control samples. The results indicated that rs1537516and rs1537515were completely linkage disequilibrium. Through case-control analisis we found that in the preterm group rs1537515and rs1537516heterozygous genotype frequencies was12.7%, significantly lower than the control group (17.9%). The results suggested that rs1537515and rs1537516heterozygous genotype may be protective factors for PTB (P=0.017, OR=0.654,95%CI=[0.47,0.91]).The results indicated that rs1537515and rs1537516associated with PTB susceptibility maybe in a linkage disequlibrium block (LD block). To further explore the two SNPs associated with PTB is due to regulate gene expression or due to the linkage with SNPs in coding regions affecting MTHFR enzyme activity, we analyzed LD of rs1537516and rs1537515to find all the linkage SNPs by Haploview4.2. Then we verified them by SNaPShot genotyping technology in order to explore the role of rs1537516and rs1537515on PTB and find other relevant SNP sites. The results showed that there was completely linkage disequilibrium between rs13306556, rs2274976, rs1537515and rs1537516, which heterozygous genotypes were protective factors (P=0.017, OR=0.65,95%CI=[0.46,0.91]). There was no linkage between rs4846048and rs1537516and rs1537515, but heterozygous genotypes of this site was predisposing factors of PTB (p=0.001, OR=1.67,95%CI=[1.24,2.25]). Further analisis showed that rs13306556and rs4846048were in noncoding region, rs2274976was in Exon11that can change glutamate into arginine. To clarify the action mechanism, functional verification of these SNPs and interaction between these SNPs will require further investigation.