Key role of Ubc5 and lysine-63 polyubiquitination in viral activation of IRF3.
The mitochondrial antiviral signaling protein (MAVS; also known as IPS-1, VISA, and CARDIF) is essential for innate immune response against RNA viruses. MAVS transduces signals from the cytosolic RIG-I-like receptors, which bind to viral RNAs. But how MAVS activates downstream transcription factors such as IRF3 to induce type-I interferons is not well understood. We have established a cell-free system in which mitochondria derived from virus-infected cells activate IRF3 in the cytosol. Fractionation of the cytosol led to the identification of Ubc5 as a ubiquitin-conjugating enzyme (E2) required for IRF3 activation. Using an inducible RNAi strategy, we demonstrate that catalytically active Ubc5 is required for IRF3 activation by viral infection. The activation of IRF3 also requires two ubiquitin-binding domains of NEMO. Furthermore, we show that replacement of endogenous ubiquitin with its K63R mutant abolishes viral activation of IRF3, demonstrating that K63 polyubiquitination plays a key role in IRF3 activation.
Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390-9148, USA.
Wenwen
W
Zeng
Ming
M
Xu
Siqi
S
Liu
Lijun
L
Sun
Zhijian J
ZJ
Chen
D000595:Amino Acid Sequence; D055162:Biocatalysis; D002458:Cell Fractionation; D006367:HeLa Cells; D006801:Humans; D051550:I-kappa B Kinase / Q000737:chemistry / Q000378:metabolism; D050838:Interferon Regulatory Factor-3 / Q000378:metabolism*; D008239:Lysine / Q000378:metabolism*; D008969:Molecular Sequence Data; D025821:Polyubiquitin / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D029082:Sendai virus / Q000502:physiology*; D044763:Ubiquitin-Conjugating Enzymes / Q000737:chemistry / Q000378:metabolism*; D054875:Ubiquitination; D014775:Virus Activation
C491731:IKBKG protein, human; C494232:IRF3 protein, human; D050838:Interferon Regulatory Factor-3; D025821:Polyubiquitin; C097308:UBE2D1 protein, human; D044763:Ubiquitin-Conjugating Enzymes; D051550:I-kappa B Kinase; D008239:Lysine