ORIGINAL ARTICLE Opposed independent effects and epistasis in the complex association of IRF5 to SLE I Ferreiro-Neira 1 , M Calaza 1 , E Alonso-Perez 1 , M Marchini 2 , R Scorza 2 , GD Sebastiani 3 , FJ Blanco 4 , I Rego 4 , R Pullmann Jr 5,17 , R Pullmann 5 , CG Kallenberg 6 , M Bijl 6 , FN Skopouli 7 , M Mavromati 7 , S Migliaresi 8 , N Barizzone 9 , S Ruzickova 10 , C Dostal 10 , RE Schmidt 11 , T Witte 11 , C Papasteriades 12 , I Kappou-Rigatou 12 , E Endreffy 13 , A Kovacs 13 , J Ordi-Ros 14 , E Balada 14 , P Carreira 15 , JJ Gomez-Reino 1,16 and A Gonzalez 1 1 Laboratorio Investigacion 2 and Rheumatology Unit, Hospital Clinico Universitario de Santiago, Santiago de Compostela, Spain; 2 Clinical Immunology, University of Milan and Fondazione IRCCS Ospedale Maggiore Policlinico, Milan, Italy; 3 UO Complessa di Reumatologia, Ospedale S. Camillo – Forlanini, Rome, Italy; 4 Servicio de Reumatologia, CH Universitario Juan Canalejo, A Corun ˜a, Spain; 5 Institute of Clinical Biochemistry, Martin Faculty Hospital, Martin, Slovakia; 6 Department of Rheumatology and Clinical Immunology, University Medical Center Groningen, Groningen, The Netherlands; 7 Pathophysiology Department, Athens University Medical School, Athens, Greece; 8 Rheumatology Unit, Second University of Naples, Caserta, Italy; 9 Department of Medical Sciences and IRCAD, Eastern Piedmont University, Novara, Italy; 10 Molecular Biology and Immunogenetics Department, Institute of Rheumatology, Prague, Czech Republic; 11 Division of Clinical Immunology, Hannover Medical School, Hannover, Germany; 12 Department of Histocompatibility and Immunology, Evangelismos Hospital, Athens, Greece; 13 Paediatrics Department, University of Szeged, Szeged, Hungary; 14 Internal Medicine, Research Laboratory in Autoimmune Diseases, Hospital Vall d’Hebron, Barcelona, Spain; 15 Rheumatology Unit, Hospital 12 de Octubre, Madrid, Spain and 16 Department of Medicine, University of Santiago de Compostela, Santiago de Compostela, Spain Genetic variation in the interferon regulatory factor 5 (IRF5) gene affects systemic lupus erythematosus (SLE) susceptibility. However, association is complex and incompletely defined. We obtained fourteen European sample collections with a total of 1383 SLE patients and 1614 controls to better define the role of the different IRF5 variants. Eleven polymorphisms were studied, including nine tag single nucleotide polymorphisms (SNPs) and two extra functional polymorphisms. Two tag SNPs showed independent and opposed associations: susceptibility (rs10488631, Po10 17 ) and protection (rs729302, Po10 6 ). Haplotype analyses showed that the susceptibility haplotype, identified by the minor allele of rs10488631, can be due to epistasis between three IRF5 functional polymorphisms. These polymorphisms determine increased mRNA expression, a splice variant with a different exon 1 and a longer proline-rich region in exon 6. This result is striking as none of the three polymorphisms had an independent effect on their own. Protection was independent of these polymorphisms and seemed to reside in the 5 0 side of the gene. In conclusion, our results help to understand the role of the IRF5 locus in SLE susceptibility by clearly separating protection from susceptibility as caused by independent polymorphisms. In addition, we have found evidence for epistasis between known functional polymorphisms for the susceptibility effect. Genes and Immunity (2007) 8, 429–438; doi:10.1038/sj.gene.6364407; published online 14 June 2007 Keywords: epistasis; haplotype analysis; systemic lupus erythematosus Introduction Systemic lupus erythematosus (SLE (MIM 152700)) is a disease that presents multiple unsolved challenges. 1 Extensive research has identified many immunological abnormalities, but their significance is still unclear. It seems that defects in T- and B-cell tolerance play a critical role as well as defects in the clearance of apoptotic cells and of immunocomplexes. Recent evidences indicate that there is also an important involvement of the innate immune system involving plasmacytoid dendritic cells (pDCs), toll-like receptors (TLRs) and type I interferons (IFN). 2 In SLE etiology, there is evidence for both an environmental and a genetic component. 3,4 This latter component accounts for the increased risk in siblings, which is about 20 times the risk in the population. However, only a few genetic factors had convincingly been identified. Here, we have explored the impact that genetic variation in the interferon regulatory factor 5 (IRF5 (MIM 607218)) locus has on SLE susceptibility. The starting point in this line of research has been a hypothesis-driven genetic study based in the important Received 3 April 2007; revised and accepted 15 May 2007; published online 14 June 2007 Correspondence: Dr A Gonzalez, Laboratorio de Investigacion 2, Hospital Clinico Universitario de Santiago, 15706-Santiago de Compostela, Travesia Choupana, A Corun ˜ a, Spain. E-mail: Antonio.Gonzalez.Martinez.Pedrayo@sergas.es 17 Current address: Gerontology Research Center, NIA, NIH, Baltimore, MD, USA. Genes and Immunity (2007) 8, 429–438 & 2007 Nature Publishing Group All rights reserved 1466-4879/07 $30.00 www.nature.com/gene