Risk SNP-Mediated Enhancer-Promoter Interaction Drives Keloid Through LncRNA DEIK

Background:Keloid is a common pathological scar formed in the repair process after skin trauma,which is characterized by excessive proliferation of fibroblasts and excessive accumulation of extracellular matrix components.Keloids showed excessive growth in dermis and subcutaneous tissue,extending beyond the scope of the original wound,and continuously invading the adjacent normal skin tissue,presenting red or dark red nodular hyperplasia.The treatment effect of keloid is often poor,and there is a high recurrence rate.Keloid not only affects the appearance of patients,but also brings itching and pain,which brings a great burden to patients’ psychology and physiology.Now most researchers believe that genetic factors are important factors of keloid onset,genome-wide association analysis(GWAS)in human chromosome 1 q41 found a single nucleotide polymorphism(SNP)rs873549 as a susceptibility loci of keloids,and the site there are people in Japan and China the crowd,but the site no in-depth study,The possible reason is that this locus is located in the intergenic region,and there is no protein-coding gene nearby.However,in fact,the susceptibility locus SNP RS873549 in this region is most significantly associated with keloid pathogenesis,suggesting that both this locus and the related strong linkage disequilibrium(LD)SNP may play an important role in keloid pathogenesis.Objectives:To investigate the function and mechanism of SNP rs873549 and the SNPs in strong LD with rs873549 in keloid.Methods:1.Bioinformatics methods such as Haploview,DHS and ENCODE were used to analyze SNP sites in strong unlinked equilibrium with RS873549,and to screen out SNP sites in cis-acting elements that may regulate the expression of nearby genes.2.Crisper-cas9 technology was used to shear the above important SNP sites,and qRT-PCR was used to analyze the expression levels of genes near the above susceptibility sites,and to analyze the regulatory effects of the above loci on nearby genes.3.The interaction between the enhancer containing rs 1348270 and the promoter of lncRNA DEIK(originally named RP11-400N13.1)In fibroblasts was investigated by chromosomal construct capture.Luciferase reporter gene was used to detect the activity of rs 1348270 enhancer.4.Knockdown experiments were used to explore the function of DEIK in keloid,,and RNA-seq was performed to investigate the mechanism that DEIK regulates collagens,POSTN and COMP expression.Results:rs 1348270,an enhancer located SNP in strong LD with rs873549,mediated looping with the promoter of DEIK.The risk variant was associated with decreased enhancer-promoter interaction and DEIK down-expression in keloid.Mechanistically,down-regulation of DEIK increased the expression of collagens and,POSTN and COMP through up-regulating BMP2.Furthermore,correlation analysis revealed that DEIK expression was inversely correlated with BMP2,POSTN and COMP expression in keloid and normal fibroblasts.Conclusions:Our findings suggested that the risk variant rs 1348270 is located in an enhancer and associated with the downregulation of DEIK in keloid,and downregulation of DEIK increases the expression of collagens,POSTN and COMP through BMP2 in keloid fibroblasts.These findings will help us more thoroughly understand the role of genetic factors play in keloid development and may lead to new strategies for keloid susceptible population screening and therapies.