Abstract
Autophagy delivers cytoplasmic constituents to autophagolysosomes and is linked to both innate and adaptive immunity. Toll-like receptor 4 (TLR4) signaling induces autophagy and recruits Beclin-1, the mammalian homolog of yeast Atg6, to the receptor complex. We found that tumor necrosis factor receptor (TNFR)-associated factor 6 (TRAF6)-mediated, Lys(63) (K63)-linked ubiquitination of Beclin-1 is critical for TLR4-triggered autophagy in macrophages. Two TRAF6-binding motifs in Beclin-1 facilitated the binding of TRAF6 and the ubiquitination of Beclin-1. Lys(117), which is strategically located in the Bcl-2 homology 3 (BH3) domain of Beclin-1, was a major site for K63-linked ubiquitination. The deubiquitinating enzyme A20 reduced the extent of K63-linked ubiquitination of Beclin-1 and limited the induction of autophagy in response to TLR signaling. Treatment of macrophages with either interferon-gamma or interleukin-1 also triggered the K63-linked ubiquitination of Beclin-1 and the formation of autophagosomes. These results indicate that the status of K63-linked ubiquitination of Beclin-1 plays a key role in regulating autophagy during inflammatory responses.
Publication types
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Research Support, N.I.H., Intramural
MeSH terms
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Animals
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Apoptosis Regulatory Proteins / genetics
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Apoptosis Regulatory Proteins / metabolism*
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Autophagy / drug effects
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Autophagy / genetics
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Autophagy / physiology*
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Beclin-1
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Blotting, Western
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Cell Line
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Cells, Cultured
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Humans
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Immunoprecipitation
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Interferon-gamma / pharmacology
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Intracellular Signaling Peptides and Proteins / genetics
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Intracellular Signaling Peptides and Proteins / metabolism*
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Lipopolysaccharides / pharmacology
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Lysine / genetics
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Lysine / metabolism
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Macrophages / cytology
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Macrophages / drug effects
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Macrophages / metabolism
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Mutation
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Nuclear Proteins / genetics
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Nuclear Proteins / metabolism
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Phagosomes / metabolism
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Protein Binding
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RNA Interference
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Signal Transduction / drug effects
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Signal Transduction / genetics
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Signal Transduction / physiology
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TNF Receptor-Associated Factor 6 / genetics
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TNF Receptor-Associated Factor 6 / metabolism*
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Toll-Like Receptor 4 / genetics
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Toll-Like Receptor 4 / metabolism*
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Ubiquitination / drug effects
Substances
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Apoptosis Regulatory Proteins
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Beclin-1
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Becn1 protein, mouse
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Intracellular Signaling Peptides and Proteins
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Lipopolysaccharides
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Nuclear Proteins
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TNF Receptor-Associated Factor 6
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Toll-Like Receptor 4
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Interferon-gamma
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Lysine