Targeted resequencing of candidate genes reveals novel variants associated with severe Behçet’s uveitis

Sang Jin Kim,Jeong-Sun Seo,Hyeong Gon Yu,Seungbok Lee,Jong-Il Kim,Changho Park

doi:10.1038/emm.2013.101

Abstract

Behçet's disease (BD) is a chronic systemic inflammatory disorder characterized by four major manifestations: recurrent uveitis, oral and genital ulcers and skin lesions. To identify some pathogenic variants associated with severe Behçet's uveitis, we used targeted and massively parallel sequencing methods to explore the genetic diversity of target regions. A solution-based target enrichment kit was designed to capture whole-exonic regions of 132 candidate genes. Using a multiplexing strategy, 32 samples from patients with a severe type of Behçet's uveitis were sequenced with a Genome Analyzer IIx. We compared the frequency of each variant with that of 59 normal Korean controls, and selected five rare and eight common single-nucleotide variants as the candidates for a replication study. The selected variants were genotyped in 61 cases and 320 controls and, as a result, two rare and seven common variants showed significant associations with severe Behçet's uveitis (P<0.05). Some of these, including rs199955684 in KIR3DL3, rs1801133 in MTHFR, rs1051790 in MICA and rs1051456 in KIR2DL4, were predicted to be damaging by either the PolyPhen-2 or SIFT prediction program. Variants on FCGR3A (rs396991) and ICAM1 (rs5498) have been previously reported as susceptibility loci of this disease, and those on IFNAR1, MTFHR and MICA also replicated the previous reports at the gene level. The KIR3DL3 and KIR2DL4 genes are novel susceptibility genes that have not been reported in association with BD. In conclusion, this study showed that target enrichment and next-generation sequencing technologies can provide valuable information on the genetic predisposition for Behçet's uveitis.

Highlights

Behcet’s disease (BD) is a systemic inflammatory disease characterized by recurrent oral aphthous ulcers, genital ulcers, skin lesions and ocular involvement.[1,2]
The etiology of BD is not understood fully, but some environmental factors are thought to trigger the immunopathogenesis of BD in people with a genetic predisposition
B51, many genetic variations have been reported to be associated with BD, including HLA-A26, MICA, TNF, TLR4 and others.[6,7,8]

Summary

INTRODUCTION

Behcet’s disease (BD) is a systemic inflammatory disease characterized by recurrent oral aphthous ulcers, genital ulcers, skin lesions and ocular involvement.[1,2] uncommon, BD may cause serious manifestation including systemic vasculitis, central nervous system involvement and sightthreatening uveitis.[1,2]. B51, many genetic variations have been reported to be associated with BD, including HLA-A26, MICA, TNF, TLR4 and others.[6,7,8] Recent genome-wide association studies have revealed an association at the IL10 and IL23R–IL12RB2 loci.[9,10]. Targeted resequencing in Behcet’s uveitis SJ Kim et al suggested the existence of several significant loci in complex diseases including BD, only a small fraction of the heritable variations have been identified. One possible explanation is that many different rare or pathogenic variants, which are difficult to find using SNP microarray experiments, contribute substantially to the genetic susceptibilities of these diseases. It has been suggested that sequencing of candidate genes would be an efficient method to investigate the contribution of previously unrecognized rare or pathogenic variants to the target disorder. We performed a replication experiment using pathogenic candidates selected from the targeted sequencing

MATERIALS AND METHODS

RESULTS

DISCUSSION