Abstract
The tripartite motif-containing protein 21 (TRIM21) plays important roles in autophagy and innate immunity. Here, we found that HECT and RLD domain containing E3 ubiquitin protein ligase 5 (HERC5), as an interferon-stimulated gene 15 (ISG15) E3 ligase, catalyzes the ISGylation of TRIM21 at the Lys260 and Lys279 residues. Moreover, IFN-β also induces TRIM21 ISGylation at multiple lysine residues, thereby enhancing its E3 ligase activity for K63-linkage-specific ubiquitination and resulting in increased levels of TRIM21 and p62 K63-linked ubiquitination. The K63-linked ubiquitination of p62 at Lys7 prevents its self-oligomerization and targeting to the autophagosome. Taken together, our study suggests that the ISGylation of TRIM21 plays a vital role in regulating self-oligomerization and localization of p62 in the autophagy induced by IFN-β.
Highlights
Type I interferons (IFNs) are ubiquitously expressed in host cells upon stimulation by pathogen-associated molecular patterns, such as nucleic acids and lipopolysaccharide, derived from viruses and bacteria
ISGylation is a reversible process catalyzed by its specific E1 activating enzyme, E2 conjugating enzyme, and E3 ligases [HECT and RLD domain containing E3 ubiquitin protein ligase 5 (HERC5) [6, 7], estrogen-responsive finger protein (EFP) [8], or human homolog of Ariadne (HHARI) [9]] in a stepwise manner, while the deISGylation from its targets is mediated by ubiquitin-specific peptidase 18 (USP18) [10] reversibly
Compared to tripartite motif-containing protein 21 (TRIM21)+/+ A549 cells, IFNβ treatment led to significant increases in the numbers of colocated puncta of p62 and autophagosomes in TRIM21−/− A549 cells (Figs. 6A, C and S7C). Consistent with these findings, reconstitution of wild-type TRIM21, but not the TRIM21(5KR) mutant, led to diminished co-location of the puncta of p62 and autophagosomes (Figs. 6B, D and S7D). These results demonstrate that ISGylation of TRIM21 induced by IFN-β plays a vital role in p62 oligomerization and autophagosome targeting by catalyzing its K63-linked ubiquitination
Summary
Type I interferons (IFNs) are ubiquitously expressed in host cells upon stimulation by pathogen-associated molecular patterns, such as nucleic acids and lipopolysaccharide, derived from viruses and bacteria. A direct relationship between ISGs and autophagy has been confirmed in a recently published study, wherein ISGylation of BECN1 induced by type I IFNs was demonstrated to be critical in regulating autophagy by competing for its K63-linked ubiquitination [11]. Inspired by this vital mechanistic insight, we aimed to identify other ISGylated proteins involved in the regulation of autophagy. Hundreds of cellular proteins were documented as being ISGylated in two proteomic profiling studies [6, 12], the results from these studies still remain to be validated, and little is known regarding the functional regulation of these ISGylated proteins
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