An Association Study Between Meiosis Related Genes HFM1, STAG3, TET1 Variant And Primary Ovarian Insufficiency

Part I. Exome Sequencing Identified the new causative gene of HFM1 for Primary Ovarian InsufficiencyObjective Primary Ovarian Insufficiency ?POI? is a kind of highly heterogeneous complex diseases, affecting 1-2% of women under the age of 40 years. Although the etiology of POI is not clear, majority of patients are idiopathic. Various data indicate that POI has a strong genetic component. Genomics method has been made in identifying causative genes on complex diseases. In this study, we will use the whole exome sequencing ?WES? in the non-syndromic POI family to identify novel candidate genes.Methods WES on lllumina platform was performed in the non-syndromic POI pedigree. Acquired reads were aligned to hg19 databases. All unreported variants following autosomal recessive heritance model were confirmed using Sanger sequencing. Then, the identified candidate gene was sequenced in 69 sporadic Chinese patients with POI.Results We identified a shared compound heterozygous mutation ?c. 1686-1 G>C and p.Ile884Ser? in two affected daughters in a meiosis-specific gene HFM1, which is very likely pathogenic variants based on an autosomal recessive inheritance mode. In addition, the entire HFM1 gene was sequenced in 69 sporadic Chinese patients with POI. Another compound heterozygous mutation ?p.Gly736Ser and c.3929₃930delinsG?was identified in a sporadic patient.Conclusions Our results indicate that meiosis-specific gene HFM1 mutations can be a cause in both familial and sporadic POI of autosomal recessive inheritance.Family-based WES could be a more powerful means in hunting novel candidate genes for POI.Part? Association analysis between meiotic gene STAG3 variation and sporadic POI in Chinese Han populationObjective STAG3 is an important meiosis-specific cohesion component, required for DNA synapsis and repair between homologous chromosomes and essential for maintaining centromere chromatid cohesion. The objection of this study was to illustrate the association between STAG3 variation and sporadic POI in Chinese Han populationMethods Mutational analysis was performed in 218 sporadic POI in Chinese Han population by target sequencing of STAG3 on lllumina platform .and confirmed in 240 healthy controls by Sanger sequencing. The sequence data were aligned with STAG3 reference sequence of hg19 to filter out the variants.Results The results indicated that we didn't find the variation of splicing site less than 30bps from the end of exons. And the variants of coding region were verified by Sanger sequencing, however, the results only supported the non-clinical significance of common variants found in NGS. We didn't find reliable results for the validation of rare mutations.Conclusions Targeted sequencing of STAG3 was performed in sporadic POI in Chinese Han population. The results showed that there was no correlation between STAG3 gene and sporadic POI. Therefore, we hypothesized that STAG3 gene mutation exist racial differences.STAG3 mutation may not be a common cause of Chinese Han POI. A large sample of further study on different ethnic population will be needed to confirm the pathogenesis of STAG3 in POI.Part ?. The correlation analysis between Meiotic Regulation gene TET1 and Primary Ovarian Insufficiency in Chinese Han populationObjective Any errors that occurring meiosis initiation or pairing, synapsis,recombination, and separation of homologous chromosomes will lead to infertility.Therefore,intensive study on the molecular regulation mechanism of meiosis will provide guidance for human reproductive health. The purpose of this study was to investigate the correlation between meiotic regulation gene TET1 and POI in Chinese Han population.Methods TET1 gene mutation screening was performed in 96 sporadic POI patients and 176 normal controls in Chinese Han population by Sanger sequencing. Meanwhile,the role of TET1 gene mutation in the pathogenesis of POI was analyzed with the literature and protein structure function prediction.Results Three novel heterozygous missense mutations in TET1 gene were identified in POI patients, namely c.832G>A/p.Glu278Ly, c. 1058C>A/p.Ala353Glu,c.l219G>C/p.Glu407Gln, which were absent in 176 controls and public databases ?the dbSNP, the 1000 Genomes Project, the NHLBI-ESP, ExAC?. The PhyloP software was used to predict the nucleotides conservation among different species, which showed that the nucleotides were weakly conserved. In addition, in silico analyses were performed to predict the functional effects of these mutations by Alamut software including SIFT,Polyphen-2 and Mutation Taster, suggesting that the variant p.Glu278Ly is predicted to probably damage the structure of the TET1 protein by SIFT.Conclusions The variation rate of TET1 gene in sporadic POI was significantly higher than that in control group and East Asia in public database, suggesting that the mutation of meiotic regulation gene TET1 may be one of the pathogenesis of POI.