Identification and Characterization of Sjogren's Syndrome-Associated Genetic Variants in the IL12A and DDX6-CXCR5 Loci

Michelle L. Joachims¹, Indra Adrianto², Audrey Johnston², John Ice², Astrid Rasmussen³, Simon Bowman⁴, David M. Lewis⁵, Lida Radfar⁶, Roald Omdal⁷, Marie Wahren-Herlenius⁸, Ilias Alevizos⁹, Torsten Witte¹⁰, Roland Jonsson^11,12, Maureen Rischmueller^13,14, Patrick M. Gaffney², Judith A. James^2,15,16, Lars Rönnblom¹⁷, Elke Theander¹⁸, Nelson L. Rhodus¹⁹, Barbara M. Segal²⁰, R. Hal Scofield^2,16,21, Courtney G. Montgomery², Xavier Mariette²², Wan-Fai Ng²³, Gunnel Nordmark²⁴, Kathy L. Sivils^2,15 and Christopher J. Lessard^2,15, ¹Arthritis and Clinical Immunology, Oklahoma Medical Research Foundation, Oklahoma City, OK, ²Arthritis and Clinical Immunology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, ³Arthritis and Clinical Immunology Research Program, Oklahoma Medical Research Foundation, USA, Oklahoma City, OK, ⁴Department of Rheumatology, University Hospitals Birmingham NHS Foundation Trust, Birmingham, United Kingdom, ⁵Department of Oral and Maxillofacial Pathology, University of Oklahoma College of Dentistry, Oklahoma City, OK, ⁶Oral Diagnosis and Radiology Department, University of Oklahoma College of Dentistry, Oklahoma City, OK, ⁷Clinical Immunology Unit, Department of Internal Medicine, Stavanger University Hospital, Stavanger, Norway, ⁸Department of Medicine, Solna, Unit of Experimental Rheumatology, Karolinska Institutet, and Karolinska University Hospital, Stockholm, Sweden, Stockholm, Sweden, ⁹Sjögren's Syndrome Clinic, National Institute of Dental and Craniofacial Research, Bethesda, MD, ¹⁰Department of Clinical Immunology and Rheumatology, Hannover Medical School, Hannover, Germany, ¹¹Broegelmann Research Laboratory, Department of Clinical Science, University of Bergen, Bergen, Norway, ¹²Department of Rheumatology, Haukeland University Hospital, Bergen, Norway, ¹³Rheumatology, Queen Elizabeth Hospital, Adelaide, Australia, ¹⁴Rheumatology, University of Adelaide, Adelaide, Australia, ¹⁵Department of Pathology, University of Oklahoma Health Sciences Center, Oklahoma City, OK, ¹⁶Department of Medicine, University of Oklahoma Health Sciences Center, Oklahoma City, OK, ¹⁷Uppsala University, Department of Medical Sciences, Rheumatology and Science for Life Laboratory, Uppsala, Sweden, ¹⁸Department of Rheumatology, Malmö University Hospital, Lund University, Sweden, Malmö, Sweden, ¹⁹Department of Diagnostic and Biological Sciences, University of Minnesota School of Dentistry, Minneapolis, MN, ²⁰Division of Rheumatology, University of Minnesota Medical School, Minneapolis, MN, ²¹US Department of Veterans Affairs Medical Center, Oklahoma City, OK, ²²Institut National de la Santé et de la Recherche Médicale, Université Paris-Sud, AP-HP, Hôpitaux Universitaires Paris-Sud, Paris, France, ²³Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom, ²⁴Rheumatology, Department of Medical Sciences, Uppsala University, Sweden, Uppsala, Sweden

Meeting: 2016 ACR/ARHP Annual Meeting

Date of first publication: September 28, 2016

Keywords: Gene Expression, genetics and genomics

Session Information

Date: Sunday, November 13, 2016

Title: Sjögren's Syndrome - Poster I: Translational Science

Session Type: ACR Poster Session A

Session Time: 9:00AM-11:00AM

Background/Purpose: Sjögren’s syndrome (SS) is an autoimmune-mediated disease with hallmark features of dry eyes/mouth and autoantibodies. Genetic susceptibility to SS involves many loci, including the previously identified association of variants at the IL12A and DDX6-CXCR5loci.

Methods: Fine mapping and imputation were used to enrich existing datasets. To improve resolution, additional data was added to total 1916 SS cases and 3684 controls. Improved imputation reference panels increased the number of variants 10-fold in the IL12A region and nearly 2-fold in the DDX6-CXCR5interval. Functional significance was studied using electrophoretic mobility shift assays (EMSA) with DNA probes encompassing the variant regions and nuclear extracts from cell lines. Bands were quantified and paired t-tests performed with replicates across multiple cell lines comprising T and B lymphocyte and monocyte lineages, as well as a non-lymphoid cell control.

Results: In the region of IL12A, rs485497 remained the most statistically significant variant. This variant links two haplotypes upstream and downstream of this polymorphism. The downstream haplotype was an eQTL for the IL12A locus, peaking at rs4680536. Variant rs4680536 and SNP rs485789 were shown to be in high linkage disequilibrium (r²=0.88). Database prediction of rs485789 revealed interaction of this variant with multiple immune-relevant transcription factors, changes in consensus transcription factor binding, and altered expression of IL12A. In the DDX6-CXCR5 region, imputation showed a pattern of association spanning beyond the length of the DDX6 coding sequence to the proximal promoter of CXCR5. The top associated variants (rs7125066 and rs7119038) did not yield evidence of regional functionality. However, 46 other variants that span the region of association were identified, and four variants predicted to affect gene binding or expression were selected for further study. DNA probes for variants rs4938572, rs12365699, rs57494551, and rs10892294 were used to test for altered protein binding in EMSA assays. Across all probe sets, three distinct nuclear protein complexes were observed in both the reference and alternate alleles, with the majority of signal in the two lower sized bands. Of the 4 probe sets, the rs4938572 variant was the only one to show a significant difference in binding between the reference and alternate alleles, with increased binding by the alternate allele. The major complexes showed significantly increased binding (p<0.05) of the alternate relative to the reference allele in 5/6 lymphocytic cell lines, but no difference in non-lymphoid cells. The most prominent differences were observed in assays using cell extracts from less differentiated cell lines.

Conclusion: Genetic variants in the IL12A and DDX6-CXCR5 loci associated with SS are likely functionally impacting genes in these loci. This study adds further evidence that genetic variants in the IL12A and DDX6-CXCR5 loci are functionally involved in the etiology of SS.

Disclosure: M. L. Joachims, None; I. Adrianto, None; A. Johnston, None; J. Ice, None; A. Rasmussen, None; S. Bowman, None; D. M. Lewis, None; L. Radfar, None; R. Omdal, None; M. Wahren-Herlenius, None; I. Alevizos, None; T. Witte, None; R. Jonsson, None; M. Rischmueller, None; P. M. Gaffney, None; J. A. James, None; L. Rönnblom, None; E. Theander, None; N. L. Rhodus, None; B. M. Segal, None; R. H. Scofield, None; C. G. Montgomery, None; X. Mariette, None; W. F. Ng, None; G. Nordmark, None; K. L. Sivils, None; C. J. Lessard, None.

To cite this abstract in AMA style:

Joachims ML, Adrianto I, Johnston A, Ice J, Rasmussen A, Bowman S, Lewis DM, Radfar L, Omdal R, Wahren-Herlenius M, Alevizos I, Witte T, Jonsson R, Rischmueller M, Gaffney PM, James JA, Rönnblom L, Theander E, Rhodus NL, Segal BM, Scofield RH, Montgomery CG, Mariette X, Ng WF, Nordmark G, Sivils KL, Lessard CJ. Identification and Characterization of Sjogren’s Syndrome-Associated Genetic Variants in the IL12A and DDX6-CXCR5 Loci [abstract]. Arthritis Rheumatol. 2016; 68 (suppl 10). https://acrabstracts.org/abstract/identification-and-characterization-of-sjogrens-syndrome-associated-genetic-variants-in-the-il12a-and-ddx6-cxcr5-loci/. Accessed .

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