Skip to Content
Merck
  • Inhibition of IRAK1 Ubiquitination Determines Glucocorticoid Sensitivity for TLR9-Induced Inflammation in Macrophages.

Inhibition of IRAK1 Ubiquitination Determines Glucocorticoid Sensitivity for TLR9-Induced Inflammation in Macrophages.

Journal of immunology (Baltimore, Md. : 1950) (2017-10-19)
Fansheng Kong, Zhiwei Liu, Viral G Jain, Kenjiro Shima, Takuji Suzuki, Louis J Muglia, Daniel T Starczynowski, Chandrashekhar Pasare, Sandip Bhattacharyya
ABSTRACT

Inflammatory responses are controlled by signaling mediators that are regulated by various posttranslational modifications. Recently, transcription-independent functions for glucocorticoids (GC) in restraining inflammation have emerged, but the underlying mechanisms are unknown. In this study, we report that GC receptor (GR)-mediated actions of GC acutely suppress TLR9-induced inflammation via inhibition of IL-1R-associated kinase 1 (IRAK1) ubiquitination. β-TrCP-IRAK1 interaction is required for K48-linked ubiquitination of IRAK1 at Lys134 and subsequent membrane-to-cytoplasm trafficking of IRAK1 interacting partners TNFR-associated factor 6 and TAK1 that facilitates NF-κB and MAPK activation. Upon costimulation of macrophages with GC and TLR9-engaging ligand, GR physically interacts with IRAK1 and interferes with protein-protein interactions between β-TrCP and IRAK1. Ablation of GR in macrophages prevents GC-dependent suppression of β-TrCP-IRAK1 interactions. This GC-mediated suppression of IRAK1 activation is unique to TLR9, as GC treatment impairs TLR9 but not TLR4 ligand-induced K48-linked IRAK1 ubiquitination and trafficking of IRAK1 interacting partners. Furthermore, mutations in IRAK1 at Lys134 prevent TLR9 ligand-induced activation of inflammatory signaling mediators and synthesis of proinflammatory cytokines to an extent comparable to GC-mediated inhibition. Collectively, these findings identify a transcription-independent, rapid, and nongenomic GC suppression of TLR9 ligand-mediated IRAK1 ubiquitination as a novel mechanism for restraining acute inflammatory reactions.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Phosphatase Inhibitor Cocktail 3, DMSO solution
Sigma-Aldrich
Phosphatase Inhibitor Cocktail 2, aqueous solution (dark coloration may develop upon storage, which does not affect the activity)
Sigma-Aldrich
Resiquimod, ≥98% (HPLC)
Sigma-Aldrich
Anti-Ubiquitin Antibody, Lys63-Specific, clone Apu3, rabbit monoclonal, clone Apu3, from rabbit
Sigma-Aldrich
Protease Inhibitor Cocktail, for use with mammalian cell and tissue extracts, DMSO solution
Sigma-Aldrich
Anti-Actin (20-33) antibody produced in rabbit, IgG fraction of antiserum, buffered aqueous solution
Sigma-Aldrich
Anti-phospho-TAK1 (Ser412) Antibody, from rabbit, purified by affinity chromatography