Background/Purpose:

RA fibroblast-like synoviocytes (FLS) display a unique aggressive phenotype with a distinct epigenetic profile marked by altered chromatin accessibility. We hypothesized that differentially utilized transcription factors (TFs) and/or promoters correlating with open chromatin regions may contribute to RA FLS behavior. We utilized ATAC-sequencing data to compare regions of open chromatin in RA FLS that are the most different when compared to OA. This filter allowed us to identify DNA sequence motifs of TFs most likely to account for RA-specific cell functions.

Methods:

ATAC-seq data from the FLS derived from RA and OA arthroplasty samples and differences corresponding to chromatin accessibility were identified using DiffBind. Loci were filtered for those present in all 11 RA samples but ≤5/10 OA samples or loci present in ≥5 RA samples but in ≤1 OA sample (and vice versa) to identify the extremes of RA-OA differences. These loci were then analyzed for DNA sequence biases using MEME (Multiple EM for Motif Elicitation). Identified motifs were compared to known TFs found in multiple databases using Tomtom. Motifs were identified with high confidence if they corresponded with known patterns in all databases and expressed in FLS.

Results:

Multiple relevant TFs motifs were associated with the extremes of RA-OA chromatin accessibility differences. Motifs for KLF4 and the E2F family of TFs were identified and among the most highly significant. The motif for the E2F family of TFs was identified in 159/575 total sites where chromatin was more open in the RA vs OA samples (MEME e = 1.2e-64, HOCOMOCO q = 2.2e-2). There were 84 genes found within 1000 kb of these 159 sites, and of these, 9 sites were found within gene bodies. RNA-seq was then utilized to identify which genes associated with E2F family binding sites in open chromatin are differentially expressed. Seven of these genes were associated with significant differential expression (CRYBG3, Metazoa_SRP, ANGPTL2, ITGB3, PMP22, GLS and U4; q < 0.05). The motif for the KLF family of TFs was identified in 18/1156 total sites where chromatin was more open in the OA vs RA samples (MEME e = 3.4e-2, TFBSshape q = 0.19). There were 18 genes identified within 1000 kb of these 1156 sites, and of these, 7 sites were found within gene bodies. None of these were differentially expressed.

Conclusion:

Using differences in chromatin access we identified TFs that are associated with neighboring genes with differential expression that distinguish RA from OA. Protein coding genes identified by this approach include proteins involved with cell adhesion that could participate in the pathogenesis of RA.

« Back to 2018 ACR/ARHP Annual Meeting

ACR Meeting Abstracts - https://acrabstracts.org/abstract/differentially-utilized-transcription-factors-and-enhancers-in-rheumatoid-arthritis-ra-fibroblast-like-synoviocytes/