Computational Analysis Of GWAS SNPs And Genes Associated With Osteoporosis And Functional Study Of Locus 3p22

Osteoporosis is a kind of bone metabolic diseasewhich is characterized by low bone mass,microstructural destruction of bone tissue,increased bone fragility and easily fracture.The heritability of bone mineral density(BMD)of approximately 50%-85%.Genome Wide Association Studies(GWAS)are common methods for studying susceptibility genes in complex diseases.Over the past decade,GWAS and meta-analysis have identified a number of susceptibility loci and single nucleotide polymorphisms(SNP)for osteoporosis.In the post-GWAS era,the greatest challenge was to elucidate the function of these SNPs and genes and their impact on the risk of osteoporosis.The aims of this study were to update osteoporosis-associated GWAS SNPs and genes,and to perform functional analysis of osteoporosis GWAS locus 3p22.1.Functional annotation of osteoporosis GWAS associated SNPs and genesA total of 43 osteoporosis-related GWAS and meta-analytical papers reported 524 GWAS SNPs with P<5×10-8.Among them,128 were intergenetic,54 located downstream of genes,43 located upstream of genes,277 in intron regions,4 in 3’ UTR regions,and 18 missense mutations.A total of 43 SNPs with potential regulatory functions with a score less than 3 were identified in the RegulomeDB database.The results of the 3D SNP annotation indicated that 3 SNPs has a score greater than 200 in the database,17 SNPs with a score between 100 and 200,and 16 SNPs with a score between 60 and 100.MiRNASNP(v2.0)predicts that three GWAS lead SNPs(rs102 6 3 64,rs884205 and rs117111740)may affect miRNA binding.Risk alleles with 6 GpG-SNPs can introduce or destroy CpG sites.SIFT identified two SNPs(rs9379084 and rs17507577)from 18 missense mutant SNPs that may be detrimental to protein function.PolyPhen identified four SNPs(rs9379084,rs17507577,rs4988321 and rs17680862)that are extremely harmful to protein function.The JASPAR database was used to analyze the differential binding between transcription factors and different alleles of SNPs.GO analysis of osteoporosis-associated genes and enrichment analysis of KEGG signaling pathways indicated that these genes were mainly enriched in cells,involved in bioregulation processes,and performed binding functions.The main enriched pathways are signaling pathways regulating pluripotent stem cells,Hippo and WNT signaling pathways.Some key node genes(hub genes)were identified by the protein interaction network constructed by STRING:CTNNB1(46),BMP2(46),BMP4(44),WNT1(28),WNT4(25),LRP5(24),TCF7L2(23),WNT2B(23)and so on.2.Functional analysis of osteoporosis GWAS locus 3p22There are 6 GWAS SNPs,gene CTNNB1 and lncRNA MSTRG.70364.1 in the locus 3p22.CTNNB1 is an important protein of WNT signaling pathway and plays an important role in bone metabolism regulation.The function of long non-coding RNA(lncRNA)MSTR.70364.1 in bone metabolism is unknown.Rs430727 can differentially bind FOXB1 to inhibit its transcriptional activity and down-regulate the expression of CTNNB1 and lncRNA MSTRG70364.1.Rs430727 can differentially bind SOX15 and promote its transcriptional activity,up-regulating the expression of CTNNB1 and lncRNA MSTRG.70364.1.However,the rs430727 could not be differentially bind with FOXA1.The lncRNA MSTRG.70364.1 located upstream of CTNNB1 can promote the expression of CTNNB1.The other two SNPs(rs401680 and rs87938)have different allelic activity but cannot differential bind to transcription factors(TCF7L2,MEF2C and EN1).