Objectives Following up the systemic lupus erythematosus (SLE) genome-wide association studies (GWAS) identification of NMNAT2 at rs2022013, we fine-mapped its 150 kb flanking regions containing NMNAT2 and SMG7 in a 15 292 case–control multi-ancestry population and tested functions of identified variants.
Methods We performed genotyping using custom array, imputation by IMPUTE 2.1.2 and allele specific functions using quantitative real-time PCR and luciferase reporter transfections. SLE peripheral blood mononuclear cells (PBMCs) were cultured with small interfering RNAs to measure antinuclear antibody (ANA) and cyto/chemokine levels in supernatants using ELISA.
Results We confirmed association at NMNAT2 in European American (EA) and Amerindian/Hispanic ancestries, and identified independent signal at SMG7 tagged by rs2702178 in EA only (p=2.4×10−8, OR=1.23 (95% CI 1.14 to 1.32)). In complete linkage disequilibrium with rs2702178, rs2275675 in the promoter region robustly associated with SMG7 mRNA levels in multiple expression quantitative trait locus (eQTL) datasets. Its risk allele was dose-dependently associated with decreased SMG7 mRNA levels in PBMCs of 86 patients with SLE and 119 controls (p=1.1×10−3 and 6.8×10−8, respectively) and conferred reduced transcription activity in transfected HEK-293 (human embryonic kidney cell line) and Raji cells (p=0.0035 and 0.0037, respectively). As a critical component in the nonsense-mediated mRNA decay pathway, SMG7 could regulate autoantigens including ribonucleoprotein (RNP) and Smith (Sm). We showed SMG7 mRNA levels in PBMCs correlated inversely with ANA titres of patients with SLE (r=−0.31, p=0.01), and SMG7 knockdown increased levels of ANA IgG and chemokine (C-C motif) ligand 19 in SLE PBMCs (p=2.0×10−5 and 2.0×10−4, respectively).
Conclusion We confirmed NMNAT2 and identified independent SMG7 association with SLE. The inverse relationship between levels of the risk allele-associated SMG7 mRNAs and ANA suggested the novel contribution of mRNA surveillance pathway to SLE pathogenesis.
- Systemic Lupus Erythematosus
- Gene Polymorphism
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Handling editor Tore K Kvien
*For the BIOLUPUS and GENLES networks.
YD and JZ contributed equally.
Contributors Conceived and designed the experiments: BPT, YD, JZ, DS, ALS, MAP; performed the experiments: YD, JZ, DS, VO, KMK, JAK; analysed the data: YD, JZ, CDL, KMK, JAK; contributed reagents/materials/analysis tools: JAJ, MAP, S-CB, MEA-R, GSA, JMA, LAC, BIF, DLK, GSG, COJ, JTM, PMG, KMS, TBN, RR-G, JDR, RHS, AMS, SAB, LMV, WS, SL, D-MC, YWS, TJV, JBH, EEB, JCE, RPK, RMC, BHH, JMG; wrote the paper: YD, JZ, VO; revised the manuscript: BPT, MAP, GSA, EEB.
Funding This work was supported by the US NIH (R01AR043814 and R21AR065626 (BPT), P01AI083194 (JBH, KMS, RPK, LAC, TJV, MEAR, COJ, BPT and PMG), P01AR049084 (RPK, JCE, EEB, GSA, JDR, RRG and MAP), R01AR064820 (EEB, MAP, RRG, JDR and LMV), P30GM103510, P30AR053483, U01AI101934 and U19AI082714 (JAJ), R01CA141700 and RC1AR058621 (MEAR), P60AR053308 and UL1TR000004 (LAC), P60AR062755 and UL1RR029882 (GSG and DLK), R01AR057172 (COJ), R01AI063274 (PMG), R01AR043274 (KMS), K08AI083790, LRPAI071651 and UL1RR024999 (TBN), R01AR43727 (MAP), K24AR002138, P60AR066464 and 1U54TR001018 (RRG), U54RR023417 (JDR), R01AR051545 and UL1RR025014 (AMS), R21AI070304 (SAB), R01AR042460, N01AR062277, P20RR020143 and R37AI024717 (JBH), P30AR048311 and P30AR055385 (EEB), R01AR033062 (RPK)), the Lupus Foundation of America (BPT), the Alliance for Lupus Research (BPT, YD, KMS, TBN, LAC, COJ and SAB), the Lupus Research Institute (TBN), the US Department of Veterans Affairs (Merit Awards; JBH and GSG), the US Department of Defense (PR094002, JBH), the Arthritis National Research Foundation (Eng Tan Scholar Award; JZ and TBN), the Arthritis Foundation (AMS and PMG), the Korea Healthcare Technology R&D Project, Ministry for Health and Welfare, Republic of Korea (HI13C2124; SCB), the Korea Health Technology R&D Project through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health & Welfare, Republic of Korea (HI13C1754; YWS), the European Science Foundation RNP (BIOLUPUS Research Network), the Wellcome Trust (TJV), Arthritis Research UK (TJV), a Kirkland Scholar Award (LAC), the Wake Forest School of Medicine Center for Public Health Genomics (CDL) and University of California Los Angeles (UCLA) Clinical and Translational Science Institute (CTSI) UL1RR033176 and UL1TR000124. Some RNA samples of healthy controls used in this study were provided by the UCLA/Center for AIDS Research Virology Core Lab which was supported by the NIH grant P30AI028697.
Competing interests None declared.
Patient consent Obtained.
Ethics approval This study was approved by the Institutional Review Boards (IRBs) or the ethnic committees at the institutions where subjects were recruited, and the overall study was approved by the IRB of the Oklahoma Medical Research Foundation..
Provenance and peer review Not commissioned; externally peer reviewed.
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