Association study of CCR6 rs3093024 with Rheumatoid Arthritis in a Pakistani cohort

Abstract

C-C Chemokine receptor 6 (CCR6), an important protein in inflammatory and immunological responses, has been previously reported to be associated with rheumatoid arthritis (RA). Therefore, in order to replicate these findings, a case-control study was conducted on 500 subjects (including 250 RA patients and 250 healthy controls) of Pakistani origin. The aim of this study was to determine the association of CCR6 rs3093024 variant with RA and identify its role in splicing events using bioinformatics tools. The clinical and demographic characteristics of the patients were collected using a well-designed questionnaire. The genotype frequencies of CCR6 rs3093024 variant were determined using tetra-primer ARMS-PCR (amplification of refractory mutation system-polymerase chain reaction) method. A significant difference was found between CCR6 rs3093024 genotype frequencies [ P = 0.0016, χ 2 = 12.915]. Similarly, a significant difference in the allele frequencies between RA patients and healthy controls was also observed [ P = 0.0003 and OR (95% CI) = 0.63 (0.49–0.80)]. The stratification of patients showed that there was a significant increase in AA genotype against AG + GG in patients [ P = 0.0014, OR (95% CI) = 2.0 (1.32–3.02)]. Furthermore, using bioinformatics analysis, it was found that CCR6 rs3093024 variant might create a potential splicing enhancer motif (SF2/ASF (IgM-BRCA1) with score of 77.92; Threshold 70.53), which might have important impact on the product of this gene. This study suggests that the A variant of CCR6 rs3093024 variant is significantly associated with RA-risk and its G variant is protective in Pakistani population but a multi-cohort large sized population study is needed to elucidate these results. Moreover, functional studies are needed to highlight the effects of this polymorphism on the function of CCR6 gene.

Characterization of Rheumatoid Arthritis Risk-Associated SNPs and Identification of Novel Therapeutic Sites Using an In-Silico Approach

“…SNPs hold a significant role in the pathogenesis of a disease. In-silico characterization underlines the possible functional significance of SNPs in both the coding and non-coding regions of a gene, for example through alteration of mRNA stability, promoter activity, and miRNA sites [14][15][16]. However, the actual underlying genetic mechanism concerning the SNPs has not been determined.…”

“…This is similarly the case in RA-associated SNPs. Previously, some studies were carried out to identify the association of SNPs with RA [14][15][16][17], but in-silico characterizations of these SNPs are scarce.…”

“…Changes were shown in eukaryotic linear motif (ELM) sites, with in a gain in intrinsic disorder and the loss of the allosteric site at position R123 in the TNFAIP3 protein. The ELM motif is a resource database which is dedicated to short linear motifs (SLiMs) [15]. These are small proteins, considered as functional modules, which play an important role in the modifications of protein sequences and protein-protein interactions (PPI) [16,17].…”

“…The SNPs located in the 5 UTR are associated with the changes in mRNA stability, binding capacity to ribosomes, and translational regulation, affecting the RNA half-life. The localization, translational efficiency, polyadenylation, stability, and binding specificity of miRNA (microRNA) may be altered by SNPs in the 3 UTR, which effects the expression patterns of genes [15]. In our study, we analyzed 11 UTR SNPs, including six 3 UTR SNPs and five 5 UTR SNPs using UTRScan, the PolymiRTS database, and MicroSNiPer.…”

“…SNPs are genetic variations that account for~0.1% differences in populations. The coding region contains about 50% SNPs, with~25% being missense and~25% being silent or synonymous [14,15]. Non-coding SNPs may change mRNA stability and promoter activity by creating or disrupting the miRNA sites, causing an altered gene expression with the consequent up-or down-regulation of a gene.…”