Glycogen synthase kinase 3β ubiquitination by TRAF6 regulates TLR3-mediated pro-inflammatory cytokine production.

Glycogen synthase kinase 3β ubiquitination by TRAF6 regulates TLR3-mediated pro-inflammatory cytokine production.

Nature communications (2015-04-02)

Ryeojin Ko, Jin Hee Park, Hyunil Ha, Yongwon Choi, Soo Young Lee

ABSTRACT

TRAF6 is critical for the production of inflammatory cytokines in various TLR-mediated signalling pathways. However, it is poorly understood how TRAF6 regulates TLR3 responses. Here we demonstrate that GSK3β interacts with TRAF6 and positively regulates the TLR3-mediated signalling. Suppression of GSK3β expression or its kinase activity drastically reduces the production of inflammatory cytokines and the induction of c-Fos by decreasing ERK and p38 phosphorylation. GSK3β physically associates with TRAF6 in a TLR3 ligand poly I:C-dependent manner. TRAF6 is determined to be a direct E3 ligase for GSK3β, and TRAF6-mediated GSK3β ubiquitination is essential for poly I:C-dependent cytokine production by promoting the TLR3 adaptor protein TRIF-assembled signalling complex.

MATERIALS

Product Number

Brand

Product Description

M8266

Sigma-Aldrich

Magnesium chloride, anhydrous, ≥98%

E9884

Sigma-Aldrich

Ethylenediaminetetraacetic acid, ACS reagent, 99.4-100.6%, powder

E2385

Supelco

Ethanol standards 10% (v/v), 10 % (v/v) in H₂O, analytical standard

Sigma-Aldrich

Ethylenediaminetetraacetic acid, purified grade, ≥98.5%, powder

EDS

Sigma-Aldrich

Ethylenediaminetetraacetic acid, anhydrous, BioUltra, ≥99% (titration)

E6758

Sigma-Aldrich

Ethylenediaminetetraacetic acid, anhydrous, crystalline, BioReagent, suitable for cell culture

M4880

Sigma-Aldrich

Magnesium chloride, BioReagent, suitable for insect cell culture, ≥97.0%

05030

Sigma-Aldrich

Sodium dodecyl sulfate solution, BioUltra, for molecular biology, 20% in H₂O

208337

Sigma-Aldrich

Magnesium chloride, powder, <200 μm

493511

Sigma-Aldrich

Ethyl alcohol, Pure, 190 proof, ACS spectrophotometric grade, 95.0%

449164

Sigma-Aldrich

Magnesium chloride, AnhydroBeads^™, −10 mesh, 99.9% trace metals basis

449172

Sigma-Aldrich

Magnesium chloride, AnhydroBeads^™, −10 mesh, 99.99% trace metals basis

71736

Sigma-Aldrich

Sodium dodecyl sulfate solution, BioUltra, for molecular biology, 10% in H₂O

431788

Sigma-Aldrich

Ethylenediaminetetraacetic acid, 99.995% trace metals basis

T7567

Sigma-Aldrich

Tributylphosphine solution, 200 mM (in N-methyl-2-pyrrolidinone), liquid

G6529

Sigma-Aldrich

L-Glutathione reduced, BioXtra, ≥98.0%

G4251

Sigma-Aldrich

L-Glutathione reduced, ≥98.0%

G6013

Sigma-Aldrich

L-Glutathione reduced, suitable for cell culture, BioReagent, ≥98.0%, powder

E8008

Sigma-Aldrich

Ethylenediaminetetraacetic acid solution, 0.02% in DPBS (0.5 mM), sterile-filtered, BioReagent, suitable for cell culture

83264

Sigma-Aldrich

HEPES buffer solution, 1 M in H₂O

459836

Sigma-Aldrich

Ethyl alcohol, Pure, 200 proof, anhydrous, ≥99.5%

V900106

Sigma-Aldrich

Ethylenediaminetetraacetic acid, Vetec^™, reagent grade, 98%

V900456

Sigma-Aldrich

L-Glutathione reduced, Vetec^™, reagent grade, ≥98%

EHU034291

Sigma-Aldrich

MISSION^® esiRNA, targeting human FOS

H4034

Sigma-Aldrich

HEPES, BioPerformance Certified, ≥99.5% (titration), suitable for cell culture

S5886

Sigma-Aldrich

Sodium chloride, BioReagent, suitable for cell culture, suitable for insect cell culture, suitable for plant cell culture, ≥99%

H6147

Sigma-Aldrich

HEPES, BioXtra, suitable for mouse embryo cell culture, ≥99.5% (titration)

S8776

Sigma-Aldrich

Sodium chloride solution, 0.9% in water, BioXtra, suitable for cell culture

L6026

Sigma-Aldrich

Sodium dodecyl sulfate, BioXtra, ≥99.0% (GC)

L5750

Sigma-Aldrich

Sodium dodecyl sulfate, 92.5-100.5% based on total alkyl sulfate content basis