TALEN-mediated enhancer knockout influences TNFAIP3 gene expression and mimics a molecular phenotype associated with systemic lupus erythematosus

Genes Immun. 2016 Apr;17(3):165-70. doi: 10.1038/gene.2016.4. Epub 2016 Jan 28.

Abstract

Linkage disequilibrium poses a major challenge to the functional characterization of specific disease-associated susceptibility variants. Precision genome-editing technologies have provided new opportunities to address this challenge. As proof of concept, we employed TALEN (transcription activation-like effector nuclease)-mediated genome editing to specifically disrupt the TT>A enhancer region to mimic candidate causal variants identified in the systemic lupus erythematosus-associated susceptibility gene, tumor necrosis factor-α-induced protein 3 (TNFAIP3), in an isogenic HEK293T cell line devoid of other linkage disequilibrium-associated variants. Targeted disruption of the TT>A enhancer impaired its interaction with the TNFAIP3 promoter by long-range DNA looping, thereby reducing TNFAIP3 gene expression. Loss of TNFAIP3 mRNA and its encoded protein, A20, impaired tumor necrosis factor-α-induced receptor-mediated downregulation of nuclear factor-κB signaling, a hallmark of autoimmunity. Results demonstrate that the TT>A enhancer variants contribute to causality and function independently of other variants to disrupt TNFAIP3 expression. Furthermore, we believe this approach can be implemented to independently examine other candidate casual variants in the future.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Down-Regulation
  • Enhancer Elements, Genetic*
  • HEK293 Cells
  • Humans
  • Lupus Erythematosus, Systemic / genetics*
  • Mutation*
  • NF-kappa B / metabolism
  • Phenotype*
  • Polymorphism, Single Nucleotide
  • Promoter Regions, Genetic
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Signal Transduction
  • Tumor Necrosis Factor alpha-Induced Protein 3 / genetics*
  • Tumor Necrosis Factor alpha-Induced Protein 3 / metabolism

Substances

  • NF-kappa B
  • RNA, Messenger
  • TNFAIP3 protein, human
  • Tumor Necrosis Factor alpha-Induced Protein 3