Next Generation Sequencing Reveals the Association of DRB3~*02:02 With Type 1 Diabetes

摘要

The primary associations of the HLA class Ⅱ genes, HLA-DRB1 and HLA-DQB1, and the class I genes, HLA-A and HLA-B, with type 1 diabetes (T1D) are well established. However, the role of polymorphism at the HLA-DRB3, HLA-DRB4, and HLA-DRB5 loci remains unclear. In two separate studies, one of 500 subjects and 500 control subjects and one of 366 DRBl~*03:01-positive samples from selected multiplex T1D families, we used Roche 454 sequencing with Conexio Genomics ASSIGN ATF 454 HLA genotyping software analysis to analyze sequence variation at these three HLA-DRB loci. Association analyses were performed on the two HLA-DRB loci haplotypes (DRB1-DRB3, -DRB4, or -DRB5). Three common HLA-DRB3 alleles (~*01:01, ~*02:02, ~*03:01) were observed. DRBl~*03:01 haplotypes carrying DRB3~*02:02 conferred a higher T1D risk than did DRBl~*03:01 haplotypes carrying DRB3~*01:01 in DRBl~*03:01y~*03:01 homozygotes with two DRB3~*01:01 alleles (odds ratio [OR] 3.4 [95% CI 1.46-8.09]), compared with those carrying one or two DRB3~*02:02 alleles (OR 25.5 [3.43-189.2]) (P = 0.033). For DRBl~*03:01/~*04:01 heterozygotes, however, the HLA-DRB3 allele did not significantly modify the T1D risk of the DRBl~*03:01 haplotype (OR 7.7 for ~*02:02; 6.8 for ~*01:01). These observations were confirmed by sequence analysis of HLA-DRB3 exon 2 in a targeted replication study of 281 informative T1D family members and 86 affected family-based association control (AFBAC) haplotypes. The frequency of DRB3~*02:02 was 42.9% in the DRBl~*03:01/~*03:01 patients and 27.6% in the DRBl~*03:01/~*04 (P = 0.005) compared with 22.6% in AFBAC DRBl~*03:01 chromosomes (P = 0.001). Analysis of T1D-associated alleles at other HLA loci (HLA-A, HLA-B, and HLA-DPB1) on DRBl~*03:01 haplotypes suggests that DRB3~*02:02 on the DRBl~*03:01 haplotype can contribute to T1D risk.