Epigenome Analysis Reveals TBX5 as a Novel Transcription Factor Involved in the Activation of Rheumatoid Arthritis Synovial Fibroblasts

Abstract

In this study, we analyzed the methylation status of human promoters in rheumatoid arthritis synovial fibroblasts (RASF). Differentially methylated genes between RASF and osteoarthritis synovial fibroblasts (OASF) were identified by methylated DNA immunoprecipitation and hybridization to human promoter tiling arrays. The methylation status was confirmed by pyrosequencing. Gene and protein expression of differentially methylated genes was evaluated with real-time PCR, Western blot, and immunohistochemistry. Chromatin immunoprecipitation was used to measure the gene promoter–associated acetylation and methylation of histones. Transcription factor–specific targets were identified with microarray and luciferase assays. We found that the transcription factor T-box transcription factor 5 (TBX5) was less methylated in rheumatoid arthritis (RA) synovium and RASF than in osteoarthritis (OA) samples. Demethylation of the TBX5 promoter in RASF and RA synovium was accompanied by higher TBX5 expression than in OASF and OA synovium. In RA synovium, TBX5 expression was primarily localized to the synovial lining. In addition, the TBX5 locus was enriched in activating chromatin marks, such as histone 4 lysine 4 trimethylation and histone acetylation, in RASF. In our functional studies, we observed that 790 genes were differentially expressed by 2–6-fold after overexpression of TBX5 in OASF. Bioinformatic analysis of these genes revealed that the chemokines IL-8, CXCL12, and CCL20 were common targets of TBX5 in OASF. Taken together, our data show that TBX5 is a novel inducer of important chemokines in RASF. Thus, we conclude that RASF contribute to the inflammatory processes operating in the pathogenesis of RA via epigenetic control of TBX5.

Cartilage damage in osteoarthritis and rheumatoid arthritis—two unequal siblings

“…Epigenetic regulation of signalling and effector molecules has been suggested as an important factor determining the permanently imprinted phenotype of these cells, [118][119][120] and epigenetic screening analyses have identified novel factors, such as the Tbox transcription factor TBX5, that contribute to their activation. 121 In addition, alterations in posttranslational protein modification, and degradation through ubiquitinationrelated mechanisms, contribute to the activation of FLS in RA. These alterations involve SUMOylation, 122-125 E3 ubiquitin-protein ligases (such as synoviolin) and changes in autophagy.…”

The Histone Modification Code in the Pathogenesis of Autoimmune Diseases

“…Signal transducer and activator of transcription 3 (STAT3), an IL-6-induced transcription factor, were shown to be associated with the MMP-1, MMP-3, and MMP-13 promoters but not the MMP-9 promoter. T-box transcription factor 5 (TBX5) expression is high in RASFs and both H3K4me3 and histone acetylation are increased in the TBX5 promoter in RASFs [85]. High IL-6 expression is associated with high levels of H3ac in the IL-6 promoter in RASFs [86].…”

The Mechanisms Underlying Chronic Inflammation in Rheumatoid Arthritis from the Perspective of the Epigenetic Landscape

“…In addition, its expression was found to be associated with specific histone markers in the promoter, such as H3K4me3 and H3K27me3. T-box transcription factor 5 (TBX5) is highly expressed in RASFs [86]. Correspondingly, active histone markers, including H3K4me3 and histone acetylation, were increased in the TBX5 promoter of RASFs.…”

“…Enforced hypermethylation of the CpG island repressed DR3 gene expression, which resulted in resistance to apoptosis in RASFs [95]. Differentially methylated genes between RASFs and OASFs were examined by methylated DNA immunoprecipitation and promoter tiling assays, which showed that TBX5 is less methylated in RASFs than in OASFs [86]. TBX5 induces the production of chemokines, such as IL-8, CXCL12, and CCL20.…”