The A Allele of the Single-Nucleotide Polymorphism rs630923 Creates a Binding Site for MEF2C Resulting in Reduced CXCR5 Promoter Activity in B-Cell Lymphoblastic Cell Lines

Abstract

Chemokine receptor CXCR5 is highly expressed in B-cells and under normal conditions is involved in their migration to specific areas of secondary lymphoid organs. B-cells are known to play an important role in various autoimmune diseases including multiple sclerosis (MS) where areas of demyelinating lesions attract B-cells by overexpressing CXCL13, the CXCR5 ligand. In this study, we aimed to determine the functional significance of single-nucleotide polymorphism rs630923 (A/C), which is located in cxcr5 gene promoter, and its common allele is associated with increased risk of MS. Using bioinformatics and pull-down assay in B-lymphoblastic cell lines, we showed that protective minor rs630923 “A” allele created functional binding site for MEF2C transcription factor. Elevated MEF2C expression in B-cells correlated with reduced activity of cxcr5 promoter containing rs630923 “A” allele. This effect that was fully neutralized by MEF2C-directed siRNA may mechanistically explain the protective role of the rs630923 minor allele in MS. Using site-directed mutagenesis of the cxcr5 gene promoter, we were unable to find any experimental evidence for the previously proposed role of NFκB transcription factors in rs630923-mediated CXCR5 promoter regulation. Thus, our results identify MEF2C as a possible mediator of protective function of the rs630923 “A” allele in MS.

A New Risk Variant for Multiple Sclerosis at 11q23.3 Locus Is Associated with Expansion of CXCR5+ Circulating Regulatory T Cells

“…However, they did not observe any difference between alleles [35]. Two years later, Mitkin et al also employed a luciferase assay to analyze the CXCR5 gene promoter activity influenced by rs630923 and observed that the minor allele (A) created a MEF2C transcription factor binding site that involved a decreased transcriptional activity in Raji cells [36]. Since rs630923 is in low LD with rs10892307 (r 2 = 0.039), the variant that best capture one of the associations in our population for the CXCR5 region, we believe that the functional effect described in Raji cells [36] must be related to other of the MS association signals of the locus.…”

The Risk G Allele of the Single-Nucleotide Polymorphism rs928413 Creates a CREB1-Binding Site That Activates IL33 Promoter in Lung Epithelial Cells

“…All PCR products were verified by sequencing; sequences of PCR primers are represented in Table S1 . Isolation of nuclear extracts, formation and immunoprecipitation of DNA-protein complexes, and the evaluation of bound DNA probes by RT-PCR were performed as described [ 72 ]. Rabbit polyclonal anti-CREB1 antibody (ab31387, Abcam, Cambridge, UK) was precipitated with pre-blocked magnetic beads (Thermo Scientific, Waltham, MA, USA).…”

The processes of homeostasis, chemotaxis and organic and inorganic response are significantly up-regulated during short-term oral mucosal cells in vitro cultivation

“…It also controls cardiac morphogenesis and myogenesis, and is likewise involved in vascular development. Enhances transcriptional activation mediated by SOX18 [16,17]. The next place in the table was placed by the TADA3 gene.…”