Transcriptional Regulation Of Peptidylarginine Deiminase Type IV: Implications For Rheumatoid Arthritis

Ali Abbas¹, Kevin Le¹, Virginia Pimmett¹, David A. Bell², Ewa Cairns² and Rodney P. DeKoter¹, ¹Microbiology and Immunology, Schulich School of Medicine and Dentistry, Western University, London, ON, Canada, ²Medicine, Division of Rheumatology, Schulich School of Medicine and Dentistry, Western University, London, ON, Canada

Meeting: 2013 ACR/ARHP Annual Meeting

Keywords: Gene Expression, PAD, Rheumatoid arthritis (RA), transcription factor and transcriptional regulation

Session Information

Title: Genetics and Genomics of Rheumatic Disease II

Session Type: Abstract Submissions (ACR)

Background/Purpose: High titers of anti-citrullinated protein/peptide antibodies (ACPA) have been detected in sera of rheumatoid arthritis (RA) patients, implicating citrullinating enzymes in the pathogenesis of RA. Peptidylarginine Deiminase Type IV (PAD4) is a member of the PAD family of citrullinating enzymes and has been linked to RA. Therefore, our aim was to determine how transcription of PAD4 is regulated in the human myeloid lineage.

Methods: The PAD4 transcription start site and promoter was located by 5’ RACE and phylogenetic comparisons of the area identified a 200 bp conserved region. Bioinformatics analysis predicted the presence of a NFκB binding site and this was tested by luciferase assays. RT-qPCR was used to quantify PAD4 expression in HL-60 cells treated with TNF-α to activate the canonical NFκB pathway. Finally, chromatin immunoprecipitation (ChIP) was used to determine NFκB enrichment at the PAD4 promoter.

Results: The human PAD4 promoter showed high biological activity. Interestingly, mutation of the predicted NFκB binding site significantly increased activity in human cell lines. PAD4 mRNA was reduced in response to TNF-α treatment. Finally, the p50 subunit of NFκB was more highly enriched than p65 at the PAD4 promoter.

Conclusion: We characterized the PAD4 promoter and demonstrated that the p50 subunit of NFκB binds to the PAD4 promoter upon NFκB activation. Our results suggest that the p50 subunit of NFκB may play a role in the repression of PAD4 transcription during inflammation.

Disclosure:

A. Abbas,
None;

K. Le,
None;

V. Pimmett,
None;

D. A. Bell,
None;

E. Cairns,
None;

R. P. DeKoter,
None.

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