Background/Purpose: Rheumatoid arthritis (RA) is a prevalent autoimmune disease in which aberrant immune attacks on joints and other tissues leads to permanent and disabling injury. Genome wide association studies (GWAS) have identified over 100 single nucleotide polymorphisms (SNPs) to the risk of RA. These variants are preferentially located in the transcriptional regulatory regions of T cells. Among these are non-coding variants in the immunologically and metabolically significant DNASE1L3/PXK/PDHB locus. However, the mechanisms by which these genetic variants contribute to disease onset are not well understood. We investigated whether non-coding variant rs73081554 regulates transcription factor binding in T cells and modulates gene expression of immune pathways.

Methods: We examined four SNPs at the DNASE1L3/PXK/PDHB locus to assess their potential impact on transcription factor binding using an electrophoretic mobility shift assay (EMSA) with nuclear extracts from Jurkat T cells and primary CD4 T cells. Additionally, we performed luciferase reporter assays in Jurkat T cells to test allele specific enhancer capabilities of the candidate SNPs. We identified potential gene targets of the SNPs tested using public eQTL data from blood samples in 31,684 healthy subjects. Gene targets were evaluated using CRISPR-Cas9 editing.

Results: EMSA data demonstrated risk (T) allele-dependent protein binding of rs73081554, an intronic SNP of the RPP14 gene, in both Jurkat T cells and primary T cells. Correspondingly, the luciferase reporter assays confirmed that rs73081554 bears risk (T) allele-specific enhancer capabilities. eQTL data revealed that the C/T variation at rs73081554 correlates most strongly (p = 7.3 x 10^-69) with RNA expression of pyruvate dehydrogenase E1 subunit beta (PDHB).

Conclusion: Our findings suggest that rs73081554 is a functional variant affecting transcription regulation in DNASE1L3/PXK/PDHB locus. PDHB is highly expressed in T cells and is a critical subunit of the pyruvate dehydrogenase complex, which converts pyruvate into mitochondrial acetyl-CoA for oxidative metabolism. This metabolic pathway is essential for the homeostatic proliferation of Treg cells, which are key to immune tolerance. This indicates that genetic susceptibility in the PDHB pathway may disrupt T cell homeostasis, contributing to RA.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/identification-of-immune-pathways-regulated-by-a-non-coding-variant-at-dnase1l3-pxk-pdhb/