A Functional Polymorphism In IFNAR1 Gene Is Associated With Susceptibility Of HFMD With EV71 Infection

ObjectiveFirstly, to explore the association between type I interferon signaling pathways and the hand,foot and mouth disease(HFMD)infection with enterovirus 71(EV71).Secondly, to explore the association between type I interferon receptor 1(IFNAR1)gene single nucleotide polymorphism(SNPs)rs2843710 and susceptibility and severity of HFMD infection with EV71 and it’s mechanism.MethodsThe mRNA profiles in Peripheral blood mononuclear cell(PBMC)from healthy donors(HD,n=10), mild EV71 HFMD(M-EV71,n=6), severe EV71 HFMD(S-EV71,n=6) were performed using gene chips. In order to verify the Chip results,we used Quantitative Real-time PCR(q RT-PCR)to detect the 4 representative gene in another cohort(n=16 respectively). At first, we selected four SNPs which are rs2843710, rs1787572, rs113181057 and rs1012334 in the gene IFNAR1 and evaluation genotyping of polymorphisms in IFNAR1 gene were associated with susceptibility and clinical phenotype of EV71 HFMD in 1196 cases and 573 controls.Then, we were evaluated the associations of the variants with EV71 HFMD using a case-control study and a mild-severe cases comparison study. In addition, we also evaluated the associations of the variants with EV71 HFMD in same gender using a case-control study. Furthermore, we validated whether the IFNAR1 gene SNP was involved in regulating the expressions of IFNAR1, IFNAR2, OAS1 and MX1 using q RT-PCR. At last, to evaluate the functional significance of promoter polymorphism, transcriptional activity of the IFNAR1 promoter variant rs2843710 was determind by promoter luciferase reporter assay.Results(1) We performed genome-wide transcriptional analysis in PBMC isolated from HD(n=10), M-EV71(n=6) and S-EV71(n=6). Clustering analysis showed that the expression of 43 related genes was clearly different in HD, M-EV71 and S-EV71 groups, which could be divided into two gene clusters according to the different clustering model. The related genes of type I IFN signaling pathways had a similar expression patterns in the same cluster.In particular, expressions of IFNAR1, IFNAR2, OAS1 and MX1 genes were significantly decreased in EV71 HFMD patients compared with HD.(2) The IFNAR1, IFNAR2, OAS1 and MX1 genes expression of PBMC were significantly decreased in EV71 HFMD patients compared with HD validated by a conventional q RT-PCR assay using another cohort(n= 16 for each group respectively). Especially, the IFNAR1, IFNAR2, OAS1 and MX1 gene expression levels were much lower in S-EV71 than those in M-EV71.(3) Among four SNPs of IFNAR1,distribution of rs2843710,rs1787572,rs113181057 and rs1012334 in EV71 HFMD cases and controls were coincident with Hardy-Weinberg equilibrium.(4) The allele frequency of rs2843710 SNP was significantly different in HD(n=573) and EV71 HFMD, and the allele G showed an increased risk to EV71 HFMD(OR=1.34, 95% CI, 1.15-1.55; P=0.0001). In genotype level, the patients with rs2843710 genotype GG showed an increased risk of EV71 HFMD in the recessive model(OR=1.33, 95% CI, 1.09-1.63; P=0.005). Genotypes CC and CG showed decreased EV71 HFMD susceptibility compared with genotype GG using an additive model(OR=0.49, 95%CI, 0.35–0.69; P <0.0001; and OR=0.58, 95%CI, 0.42–0.82, P= 0.002).(5) In male population, the rs2843710 allele G frequency was significantly higher in EV71 HFMD cases than that in controls in the multiplicative model(OR=1.57, 95% CI, 1.29-1.90; P<0.0001). In genotype level, the patients with rs2843710 genotype GG increased risk of EV71 HFMD in the recessive model(OR = 1.63, 95% CI, 1.25 2.11; P=0.0002). In contrast, rs2843710 showed no association with susceptibility to EV71 HFMD in female population.(6) The frequency of the rs2843710 allele G was significantly higher in S-EV71 patients than that in M-EV71 patients(OR=1.21, 95% CI, 1.02-1.44; P=0.028).(7) The expressions of IFNAR1, OAS1 and MX1 were much lower in genotype GG than that in genotypes CC or CG, while rs2843710 SNP did not affect the IFNAR2 gene expression,in PBMC of HFMD infection with EV71.(8) The promoter luciferase reporter assay showed that after EV71 infection, allele G of rs2843710 significantly weaker transcriptional activity than allele C, while no difference was observed between both alleles without infection.Conclusions(1) EV71 HFMD was associated with the decrease of type I IFN related genes levels in PBMC;(2) Rs2843710 of IFNAR1 was associated with susceptibility and severity of EV71 HFMD;(3) Rs2843710 of IFNAR1 influenced IFNAR1, OAS1 and MX1 gene expression.(4) Rs2843710 of IFNAR1 affects the IFNAR1 promoter transcriptional activity.