Eukaryotic elongation factor 2 controls TNF-α translation in LPS-induced hepatitis
Bárbara González-Terán, … , Roger J. Davis, Guadalupe Sabio
Bárbara González-Terán, … , Roger J. Davis, Guadalupe Sabio
Published December 3, 2012
Citation Information: J Clin Invest. 2013;123(1):164-178. https://doi.org/10.1172/JCI65124.
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Research Article Inflammation

Eukaryotic elongation factor 2 controls TNF-α translation in LPS-induced hepatitis

Abstract

Bacterial LPS (endotoxin) has been implicated in the pathogenesis of acute liver disease through its induction of the proinflammatory cytokine TNF-α. TNF-α is a key determinant of the outcome in a well-established mouse model of acute liver failure during septic shock. One possible mechanism for regulating TNF-α expression is through the control of protein elongation during translation, which would allow rapid cell adaptation to physiological changes. However, the regulation of translational elongation is poorly understood. We found that expression of p38γ/δ MAPK proteins is required for the elongation of nascent TNF-α protein in macrophages. The MKK3/6-p38γ/δ pathway mediated an inhibitory phosphorylation of eukaryotic elongation factor 2 (eEF2) kinase, which in turn promoted eEF2 activation (dephosphorylation) and subsequent TNF-α elongation. These results identify a new signaling pathway that regulates TNF-α production in LPS-induced liver damage and suggest potential cell-specific therapeutic targets for liver diseases in which TNF-α production is involved.

Authors

Bárbara González-Terán, José R. Cortés, Elisa Manieri, Nuria Matesanz, Ángeles Verdugo, María E. Rodríguez, Águeda González-Rodríguez, Ángela Valverde, Pilar Martín, Roger J. Davis, Guadalupe Sabio

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Figure 7

eEF2 controls TNF-α elongation, and its gene silencing protects against LPS-induced liver damage.

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eEF2 controls TNF-α elongation, and its gene silencing protects against ...
(A) BM-derived macrophages from Lyzs-cre and p38γ/δLyz-KO mice were transduced with GFP+ lentivirus expressing eEf2 shRNA (shEF2) or with empty vector (shΦ). ELISA of TNF-α and IL-6 in culture supernatants was performed 16 hours after LPS (10 μg/ml) stimulation. One-way ANOVA coupled to Bonferroni’s post tests (n = 6). (B–F) WT mice were i.v. injected with shEF2 or shΦ GFP+ lentiviral vectors. After 7 days, mice were injected with D-gal+LPS or saline. Data are means ± SD. *P < 0.05; **P < 0.01; ***P < 0.001 (2-way ANOVA coupled to Bonferroni’s post tests). (B) ELISA of TNF-α and IL-6 in mouse serum 6 hours after treatment (n = 6). (C) Representative livers and H&E-stained sections after removal at 6 hours after injection The chart presents hemorrhagic area as a percentage of the total area (n = 6). Scale bar: 50 μm. (D) Serum transaminase activity at 6 hours after injection (n = 6). (E) Immunoblot analysis of liver extracts (n = 6). (F) Mouse survival after D-gal+LPS injection (n = 10). Survival curves were created with the Kaplan-Meier method and compared by log-rank (Mantel-Cox) test. (G) The MKK3/6-p38γ/δ-eEF2K pathway is necessary for LPS-induced hepatitis. LPS induces leukocyte egress from BM and cytokine/chemokine production in peripheral blood and by Kupffer cells. This leads to massive leukocyte infiltration in the liver and hepatocellular injury. TNF-α plays a dominant role in LPS-induced liver injury. Activation of the MKK3/6-p38γ/δ-eEF2K pathway in macrophages/Kupffer cells controls TNF-α production at the mRNA elongation level in acute inflammatory responses to LPS.