Citation
- Authors: Di Q. et al.
- Year: 2022
- Journal: Autophagy 1-13
- Applications: in vitro / in vivo / siRNA / in vivo-jetPEI, jetPRIME
- Cell types:
- Name: Mouse peritoneal macrophages
Description: Mouse primary peritoneal macrophage - Name: THP-1
Description: Human acute monocytic leukaemia cells
Known as: THP1, THP 1
- Name: Mouse peritoneal macrophages
Method
In vivo siRNA transfection was carried out using in vivo-jetPEI (PolyPlus Transferion, 101,000,030), according to the manufacturer’s instructions. Briefly, mice were intraperitoneally injected with transfection mixture consisting of Usp22-siRNA or control-siRNA dissolved in in vivo jetPEI with an N:P ratio of 7. The interfering efficacy was determined by quantitative PCR (qPCR) and western blotting.
The Usp22 and control-siRNA oligo complex with in vivo-jetPEI significantly reduced the endogenous Usp22 mRNA level in the peritoneal cavity.
in vitro: The siRNAs were synthesized and transfected with jetPRIME (PolyPlus Transferion, 101,000,046), according to the manufacturer’s instructions. USP22 was efficiently downregulated in mouse peritoneal macrophages and human THP-1 cells by specific small interfering RNA (siRNA) (Fig. S2A and B).
Abstract
The NLRP3 inflammasome is involved in a diverse range of inflammatory diseases. The activation of inflammasomes must be tightly regulated to prevent excessive inflammation, and the protein ubiquitination system is reported to be one of the ways in which inflammasome activation is regulated. However, the deubiquitination regulatory mechanisms of inflammasome activation remain elusive. Here, we demonstrated that USP22 (ubiquitin specific peptidase 22) promotes NLRP3 degradation and inhibits NLRP3 inflammasome activation. USP22 deficiency or in vivo silencing significantly increases alum-induced peritonitis and lipopolysaccharide-induced systemic inflammation. Mechanistically, USP22 inhibits NLRP3 inflammasome activation via the promotion of ATG5-mediated macroautophagy/autophagy. USP22 stabilizes ATG5 via decreasing K27- and K48-linked ubiquitination of ATG5 at the Lys118 site. Taken together, these findings reveal the role USP22 plays in the regulation of NLRP3 inflammasome activation and suggest a potential therapeutic target to treat NLRP3 inflammasome-related diseases.