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Arthritis Res Ther. 2012; 14(Suppl 3): A5.
Published online Sep 27, 2012. doi:  10.1186/ar3939
PMCID: PMC3467482
MicroRNA-3148 modulates differential gene expression of the SLE-associated TLR7 variant
Y Deng,1 J Zhao,1 D Sakurai,1 KM Kaufman,2,3 JC Edberg,4 RP Kimberly,4 DL Kamen,5 GS Gilkeson,5 CO Jacob,6 RH Scofield,7,8,9 CD Langefeld,10 JA Kelly,7 ME Alarcón-Riquelme, JB Harley,2,3 TJ Vyse,12 BI Freedman,13 PM Gaffney,7 KM Sivils,7 JA James,7,8 TB Niewold,14 RM Cantor,1 W Chen,1 BH Hahn,1 EE Brown, and BP Tsaocorresponding author1, BIOLUPUS and GENLES Networks , PROFILE
1University of California, Los Angeles, CA, USA
2Center for Autoimmune Genomics & Etiology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH, USA
3US Department of Veterans Affairs Medical Center, Cincinnati, OH, USA
4University of Alabama at Birmingham, Birmingham, AL, USA
5Medical University of South Carolina, Charleston, SC, USA
6Keck School of Medicine, University of Southern California, Los Angeles, CA, USA
7Arthritis & Clinical Immunology Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, USA
8University of Oklahoma Health Sciences Center, Oklahoma City, OK, USA
9US Department of Veterans Affairs Medical Center, Oklahoma City, OK, USA
10Wake Forest University Health Sciences, Wake Forest, NC, USA
11Centro de Genómica e Investigación Oncológica (GENYO), Pfizer-Universidad de Granada-Junta de Andalucia, Granada, Spain
12King's College London, UK
13Wake Forest School of Medicine, Winston-Salem, NC, USA
14Gwen Knapp Center for Lupus and Immunology Research, University of Chicago, IL, USA
corresponding authorCorresponding author.
Supplement
Lupus 2012: New targets, new approaches
Peter E Lipsky, John M Esdaile, Matthew H Liang and Paul R Fortin
Conference
Lupus 2012: New targets, new approaches
27-30 September 2012
Whistler, Canada
Background
We identified the G allele of TLR7 3'-UTR SNP (rs3853839) associated with increased TLR7 transcripts, a more pronounced IFN signature and risk for SLE in 9,274 Eastern Asians (Pcombined = 6.5 × 10-10) [1]. The current study sought replication of SLE-associated SNP(s) in non-Asian ancestries and explored molecular mechanisms underlying an identified gene variant that affects TLR7 expression.
Methods
We conducted genotyping, imputation and association for 98 to 116 SNPs (varying among different ancestries) covering 80 kb of TLR7-TLR8 in European Americans (EA), African Americans (AA) and Hispanics enriched for the Amerindian-European admixture (HS). Haplotype-based conditional testing was conducted to distinguish independent association signals. Mantel-Haenszel testing was used in transancestral meta-analysis. Association of genotypes with TLR7 expression was examined using RT-PCR, flow cytometry and reporter assays. Pyrosequencing was used to measure allelic variations in TLR7 transcript levels.
The rs3853839 was confirmed as the only variant within TLR7-TLR8 exhibiting consistent and independent association with SLE in our transancestral fine-mapping (Pmeta = 7.5 × 10-11, OR (95% CI) = 1.24 (1.18 to 1.34)) in 13,339 subjects of EA (3,936 cases vs. 3,491 controls), AA (1,679 vs. 1,934) and HS (1,492 vs. 807) ancestries. PBMCs from normal G-allele carriers exhibited elevated levels of TLR7 mRNA (P = 0.01 in men and P = 0.02 in women) and protein (P = 0.009 in men and P = 0.038 in women). PBMCs from heterozygotes exhibited higher G/C allele ratios of TLR7 transcripts 4 hours after incubation with actinomycin D (inhibitor of transcription initiation) (P = 0.04), indicating slower degradation of G allele-containing transcript. The nonrisk allele, but not the risk allele, was predicted to match microRNA-3148 (miR-3148) at the second base in the binding site. Transcript levels of miR-3148 and TLR7 were inversely correlated in PBMCs from 16 SLE patients and 21 controls (R2 = 0.255, P = 0.001), suggesting miR-3148 modulating TLR7 expression. Overexpression of miR-3148 via transfection into HEK 293 cells led to a more than twofold reduction in luciferase activity driven by the TLR7 3'-UTR segment containing the nonrisk allele than that containing the risk allele (P = 0.001).
Conclusion
We identified and confirmed a genome-wide significant association between rs3853839 and SLE susceptibility in 22,613 subjects of Eastern Asian, EA, AA and HS ancestries (Pmeta = 6.4 × 10-19, OR (95% CI) = 1.26 (1.20 to 1.32)). Reduced modulation by miR-3148 confers slower degradation of the risk allele containing TLR7 transcript, resulting in elevated levels of gene products and a more robust type I IFN signature.
References
  • Shen N, Fu Q, Deng Y, Qian X, Zhao J, Kaufman KM, Wu YL, Yu CY, Tang Y, Chen JY, Yang W, Wong M, Kawasaki A, Tsuchiya N, Sumida T, Kawaguchi Y, Howe HS, Mok MY, Bang SY, Liu FL, Chang DM, Takasaki Y, Hashimoto H, Harley JB, Guthridge JM, Grossman JM, Cantor RM, Song YW, Bae SC, Chen S. et al. Sex-specific association of X-linked Toll-like receptor 7 (TLR7) with male systemic lupus erythematosus. Proc Natl Acad Sci USA. 2010;107:15838–15843. doi: 10.1073/pnas.1001337107. [PubMed] [Cross Ref]
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