Citation

  • Authors: Xu J. et al.
  • Year: 2021
  • Journal: Cell Rep 37 109926
  • Applications: in vitro / DNA, siRNA / INTERFERin, jetPEI, jetPRIME
  • Cell types:
    1. Name: A549
      Description: Human lung carcinoma cells, type II pneumocytes
      Known as: A-549
    2. Name: HEK-293T
      Description: Human embryonic kidney Fibroblast
      Known as: HEK293T, 293T
    3. Name: MEF
      Description: Murine embryonic fibroblast cells 
    4. Name: Mouse peritoneal macrophages
      Description: Mouse primary peritoneal macrophage

Method

4 × 105 Cells were seeded into 6-well plate. The next day, 20 nM of indicated gene-targeting siRNA was transfected into cells using INTERFERin reagent (Polyplus Transfection) according to the manufacturer’s instruction 8 h later followed by changing fresh culture medium. 24 h later for lncRNA interference or 48 h later for protein interference, cells were proceeded to indicated assay. The plasmids were transfected into indicated cells with jetPEI reagent (Polyplus Transfection) according to the manufacturer’s instructions 8 h later followed by changing fresh culture medium. 24-36 h after transfection, the cells were used to perform the indicated assay.

Abstract

Interferon regulatory factor 3 (IRF3) is an essential transductor for initiation of many immune responses. Here, we show that lncRNA-ISIR directly binds IRF3 to promote its phosphorylation, dimerization, and nuclear translocation, along with enhanced target gene productions. In vivo lncRNA-ISIR deficiency results in reduced IFN production, uncontrolled viral replication, and increased mortality. The human homolog, AK131315, also binds IRF3 and promotes its activation. More important, AK131315 expression is positively correlated with type I interferon (IFN-I) level and severity in patients with lupus. Mechanistically, in resting cells, IRF3 is bound to suppressor protein Flightless-1 (Fli-1), which keeps its inactive state. Upon infection, IFN-I-induced lncRNA-ISIR binds IRF3 at DNA-binding domain in cytoplasm and removes Fli-1's association from IRF3, consequently facilitating IRF3 activation. Our results demonstrate that IFN-I-inducible lncRNA-ISIR feedback strengthens IRF3 activation by removing suppressive Fli-1 in immune responses, revealing a method of lncRNA-mediated modulation of transcription factor (TF) activation.

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