| Objective:To search the possible pathogenic genes and gene mutation sites in type 2diabetic pedigree.Methods:The family in which at least two generations of immediate family members were diagnosed as diabetes was chooesd.There were twenty family members in total and the proband was a female patient with diabetes for 10 years in the second generation.Peripheral venous blood was collected,and blood cells and plasma were separated.Blood cell DNA was extracted by using the whole blood genomic DNA rapid extraction kit,and then genome-wide exon sequencing and bioinformatics data analysis were performed to screen out the possible pathogenic genes and mutation sites of pedigree.Results:Among the members of the diabetes pedigree,the first generation was the proband's mother who was diagnosed as diabetes,and the father died.There were 5 persons in the second generation of the immediate family,4 of whom were diagnosed as diabetes.The third generation consists of 10 people,none of whom have been diagnosed as diabetes.The second generation with normal blood glucose were choosed as the normal control.Considering that all the third-generation immediate family members are young,though they have not been diagnosed as diabetes at present,they may also carry the pathogenic genes and mutation sites of this family and may also develop diabetes a few years later.Therefore,taking the sequencing results of the third generation into account,the candidate genes should be allowed to appear no more than 50%in third-generation family members.485 DNA mutations,including 429 single nucleotide(SNP)mutations,insertions and deletions of 56(Indel)mutations were found by using the whole exons genome sequencing and bioinformatics data analysisin this family.These DNA mutations existed in 5 patients with diabetes but did not exist in the 5 normal controls and the frequencies in the third generation of immediate family members were not more than 50%.The significant differences of these sites were analyzed,then 39 SNP mutation sites and 32 Indel mutation sites with medium and high biological functions were screened out.Because their MAF were more than 0.001,those SNPs and Indel siteswere excluded.Then 15 significant SNP mutation sites and 1 Indel mutation site were found.By consulting the literatures and locating the genes corresponding to the above 16 mutations into the KEGG pathway,EPHX3 and FHOD1 genes were finally screened out.Among them,there was a complex genetic interaction between EPHX3 gene and PIK3CB gene which was in KEGG pathway.Although there was no genetic interaction between FHOD1 gene and those genes in KEGG pathway,the FHOD1 gene might alsolead to T2DMaccording to literatures.All the diabetic patients in this study had heterozygous mutation of EPHX3 gene rs760409070[c.778 G>A(p.glu2601ys)]and heterozygous mutation of FHOD1 gene rs 142023201[c.1493 A>G(p.asn498ser)].The mutation of EPHX3 gene rs760409070 reducedthe function of its coding protein,inhibitedthe expression of PIK3CB gene that was involved in glucose transporter,glycogen synthesis,glycolysis and sugar dysplasia in the PI3K/AKT signaling pathway.Thismutation inhibited GLUT4,which transported glucose,GSK-3,which promoteed glycogen synthesis,and FOXO1,which inhibited gluconeogenesis,leading to insulin resistance and impaired glucose utilization,and ultimately to diabetes.The rs 142023201 mutation of FHOD1 gene inhibited the effect of the encoding product FHOD1 protein on GLUT4-containing vesicles of IRAP,down-regulated the function of GLUT4 uptake and glucose transport,and increased the body's blood glucose level,leading to diabetesultimately.In summary,the mutations of EPHX3 gene rs760409070[c.778 G>a(p.glu2601ys)]and FHOD1 gene rs142023201[c.1493 A>G(p.asn498ser)]may be the pathogenic genes and mutation sites of type 2 diabetes pedigreein this study.Conclusion:Mutation of EPHX3 gene rs760409070[c.778 G>A(p.glu2601ys)]and FHOD1 gene rs 142023201[c.1493 A>G(p.asn498ser)]may be the pathogenic genes and mutation sites ofT2DM amongthe family members. |
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