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Systemic lupus erythematosus
KLF2 enhancer variant rs4808485 increases lupus risk by modulating inflammasome machinery and cellular homoeostasis
  1. Manish Kumar Singh1,
  2. Harikrishna Reddy Rallabandi1,
  3. Xu-Jie Zhou2,3,4,
  4. Yuan-Yuan Qi5,
  5. Zhan-Zheng Zhao5,
  6. Ting Gan2,3,
  7. Hong Zhang2,3,
  8. Loren L Looger6,
  9. Swapan K Nath1
  1. 1Arthritis and Clinical Immunology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, Oklahoma, USA
  2. 2Renal Division, Peking University Institute of Nephrology, Peking University First Hospital, Beijing, China
  3. 3Key Laboratory of Renal Disease, Ministry of Health of China, Beijing, China
  4. 4Key Laboratory of Chronic Kidney Disease Prevention and Treatment (Peking University), Ministry of Education, Beijing, China
  5. 5Department of Nephrology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan Province, China
  6. 6Howard Hughes Medical Institute, Department of Neurosciences, University of California San Diego, La Jolla, California, USA
  1. Correspondence to Dr Swapan K Nath, Arthritis and Clinical Immunology, Oklahoma Medical Research Foundation, Oklahoma City, Oklahoma, USA; Swapan-Nath{at}omrf.org

Abstract

Objective A recent genome-wide association study linked KLF2 as a novel Asian-specific locus for systemic lupus erythematosus (SLE) susceptibility. However, the underlying causal functional variant(s), cognate target gene(s) and genetic mechanisms associated with SLE risk are unknown.

Methods We used bioinformatics to prioritise likely functional variants and validated the best candidate with diverse experimental techniques, including genome editing. Gene expression was compared between healthy controls (HCs) and patients with SLE with or without lupus nephritis (LN+, LN−).

Results Through bioinformatics and expression quantitative trait locus analyses, we prioritised rs4808485 in active chromatin, predicted to modulate KLF2 expression. Luciferase reporter assays and chromatin immunoprecipitation-qPCR demonstrated differential allele-specific enhancer activity and binding of active histone marks (H3K27ac, H3K4me3 and H3K4me1), Pol II, CTCF, P300 and the transcription factor PARP1. Chromosome conformation capture-qPCR revealed long-range chromatin interactions between rs4808485 and the KLF2 promoter. These were directly validated by CRISPR-based genetic and epigenetic editing in Jurkat and lymphoblastoid cells. Deleting the rs4808485 enhancer in Jurkat (KO) cells disrupted NLRP3 inflammasome machinery by reducing KLF2 and increasing CASPASE1, IL-1β and GSDMD levels. Knockout cells also exhibited higher proliferation and cell-cycle progression than wild type. RNA-seq validated interplay between KLF2 and inflammasome machinery in HC, LN+ and LN−.

Conclusions We demonstrate how rs4808485 modulates the inflammasome and cellular homoeostasis through regulating KLF2 expression. This establishes mechanistic connections between rs4808485 and SLE susceptibility.

  • Lupus Erythematosus, Systemic
  • Polymorphism, Genetic
  • Immune System Diseases
  • Risk Factors

Data availability statement

Data are available on reasonable request.

https://doi.org/10.1136/ard-2023-224953

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    Data availability statement

    Data are available on reasonable request.

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    Footnotes

    • Handling editor Josef S Smolen

    • Contributors MKS performed the experiments, interpreted results and participated in manuscript writing. HRR performed some experiments and interpreted results. X-JZ, Z-ZZ, TG and HZ collected and analysed the clinical information for the RNA-seq experiment. Y-YQ collected, analysed and interpreted the clinical information for the RNA-seq experiment. LLL provided intellectual input and assisted with the preparation of the manuscript. SKN, the guarantor for the manuscript, conceived, designed and supervised the study, and finalised the manuscript. All authors contributed and finally approved the current manuscript.

    • Funding Research reported in this publication was supported by National Institutes of Health grants R21AI168943 and R01AI172255 (SKN) as well as other supporting grants from the Presbyterian Health Foundation.

    • Disclaimer The content is solely the responsibility of the authors and does not necessarily reflect the official views of the National Institutes of Health.

    • Competing interests None declared.

    • Patient and public involvement Patients and/or the public were not involved in the design, or conduct, or reporting, or dissemination plans of this research.

    • Provenance and peer review Not commissioned; externally peer reviewed.

    • Supplemental material This content has been supplied by the author(s). It has not been vetted by BMJ Publishing Group Limited (BMJ) and may not have been peer-reviewed. Any opinions or recommendations discussed are solely those of the author(s) and are not endorsed by BMJ. BMJ disclaims all liability and responsibility arising from any reliance placed on the content. Where the content includes any translated material, BMJ does not warrant the accuracy and reliability of the translations (including but not limited to local regulations, clinical guidelines, terminology, drug names and drug dosages), and is not responsible for any error and/or omissions arising from translation and adaptation or otherwise.