OP0028 Independent and Interactive Effects of Interferon (IFN)-Alpha and the Lupus Risk Haplotype HLA-DRB1*03:01 on Gene Expression in Ex Vivo B Cells
Authors: C. Duarte, T. Vyse, L. Boteva, M. Fernando
Journal: Annals of the Rheumatic Diseases (2015)

<h3>Background</h3> Systemic lupus erythematosus (SLE) is a complex genetic autoimmune disease characterised by B cell hyperactivity and up-regulation of type I IFN. The <i>DRB1*03:01</i> extended haplotype confers the greatest genetic risk for SLE in Europeans, but the molecular mechanisms underlying this association are unclear. Both coding and non-coding genetic variants arising from this haplotype may contribute to disease risk, implicating a role for gene regulation. Genetic effects on gene expression are context-specific, depending on factors such as cell type and cell state. <h3>Objectives</h3> To investigate the effect of (i) IFN-α stimulation, (ii) <i>DRB1*03:01</i> haplotypes and (iii) the haplotype-environment interaction between <i>DRB1*03:01</i> and IFN-α on differential gene expression in <i>ex vivo</i> B cells. <h3>Methods</h3> RNA was extracted from <i>ex vivo</i> CD19+ B cells from 50 healthy European women harbouring <i>DRB1*03:01</i> homozygous (n=17), heterozygous (n=3) and non-<i>DRB1*03:01</i> (n=30) haplotypes. Gene expression from cells at rest (n=49) and after stimulation with IFN-α for 6 hours (n=33) was quantified using the Affymetrix Human Exon 1.0 ST array. The individual and interactive effects of IFN-α stimulation and the <i>DRB1*03:01</i> haplotype on gene expression were analysed using a two-way ANOVA (FDR 5%) using Partek Genomics Suite. Pathway and disease enrichment were analysed using Ingenuity Pathway Analysis. <h3>Results</h3> 6,906 out of 15,468 genes are significantly differentially expressed in IFN-α-treated compared to resting cells, of which 1,263 have fold changes &gt;2. As expected, the data is enriched for the IFN-α pathway (P=3.4x10<sup>-2</sup>) and B cell receptor signalling genes (P=2.1x10<sup>-12</sup>). Approximately 60% of SLE- and rheumatoid arthritis-associated genes from GWAS are significantly enriched in this dataset (P<sub>SLE</sub>=4.4x10<sup>-2</sup>, P<sub>RA</sub>=6.1x10<sup>-6</sup>), in comparison to the non-autoimmune disease osteoarthritis (20%; P<sub>OA</sub>=0.6). Interestingly, IFN-α affects SLE-associated genes outside canonical type I IFN and B cell signalling pathways, such as <i>ITGAM</i> (P=1.9x10<sup>-10</sup>, ↓) and <i>PTPN22</i> (P=4.5x10<sup>-10</sup>, ↓). Genes implicated in monogenic SLE such as <i>TREX1</i> and <i>DNASE1L</i> (DNA repair) and <i>C1R</i> (complement pathway) are also differentially expressed on IFN-α stimulation. The only gene significantly differentially expressed between <i>DRB1*03:01</i> and non-<i>DRB1*03:01</i> samples is <i>BTN3A2</i>, which is downregulated in <i>DRB1*03:01</i> haplotypes in both resting (P=2.3x10<sup>-2</sup>) and IFN-α-treated cells (P=1.2x10<sup>-2</sup>). Furthermore, <i>BTN3A2</i> levels differ between <i>DRB1*03:01</i> and non-<i>DRB1*03:01</i> individuals in response to IFN-α stimulation, implicating haplotype-environment interactions (P<sub>int</sub>=1.4x10<sup>-7</sup>). <h3>Conclusions</h3> Stimulation of B cells with IFN-α influences the expression of genes associated with complex and monogenic forms of SLE not known to be involved in type I IFN or B cell signalling pathways. Individuals harbouring <i>DRB1*03:01</i> haplotypes demonstrate downregulation of <i>BTN3A2</i>, implicating effects independent of HLA alleles for this risk haplotype. Additionally, we show that IFN-α and <i>DRB1*03:01</i> interactively downregulate <i>BTN3A2</i>, shedding some light into the mechanism behind the known association of IFN-α exposure with SLE risk. Investigating the effects of disease-associated haplotypes on gene expression in relevant contexts such as IFN-α stimulation may help in resolving genetic associations and identifying candidate genes underlying susceptibility to complex diseases. <h3>Disclosure of Interest</h3> None declared