Regulation of Eotaxin Gene Expression by TNF-α and IL-4 Through mRNA Stabilization: Involvement of the RNA-Binding Protein HuR

Abstract

During inflammatory responses, a major posttranscriptional regulation of early response and inflammatory gene expression occurs through modulation of mRNA turnover. We report that two potent inducers of the CC chemokine eotaxin, TNF-α and IL-4, regulate its production in airway epithelial cells by increasing eotaxin mRNA stability. In experiments using the transcriptional inhibitor actinomycin D, eotaxin mRNA half-life was significantly prolonged by cell stimulation with TNF-α or IL-4, with the combination of the two cytokines being the most effective in extending the mRNA half-life. Involvement of the eotaxin 3′ untranslated region in the mRNA-stabilizing effect was tested by transient transfection of a construct expressing a chimeric transcript carrying a serum-inducible β-globin reporter linked to the eotaxin 3′ untranslated region. The half-life of the chimeric mRNA was markedly increased in cells stimulated with TNF-α and IL-4. Evidence that the mRNA-stabilizing protein HuR participated in the cytokine effect was obtained: first, HuR presence in the cytoplasm, believed to be required for HuR-mediated mRNA stabilization, increased in both transformed (BEAS-2B cell line) and primary bronchial epithelial cells following treatment with TNF-α and IL-4. Second, endogenous eotaxin mRNA was found to bind to HuR in vivo, as detected by immunoprecipitation of HuR-containing messenger ribonucleoprotein complexes followed by real-time RT-PCR analysis; such association increased after cell treatment with TNF-α and IL-4. Third, overexpression of HuR in BEAS-2B cells significantly increased the expression of eotaxin mRNA and protein. Our findings implicate mRNA stabilization in the cytokine-mediated increase in eotaxin expression and strongly suggest a role for HuR in this effect.

Activation of the Nrf2/antioxidant response pathway increases IL-8 expression

“…Although transcriptional regulation of IL-8 has been clearly demonstrated and is thought to be the main mechanism of gene expression [12,23], there is also evidence for regulation by mRNA stabilization [24,25]. Although the factors responsible for enhanced RNA stability after Nrf2 activation are not currently known, we postulate that an Nrf2-dependent RNA-binding protein may directly stabilize IL-8 mRNA, similar to the effect of the protein HuR on the RNA stability of other chemokine genes [26].…”

Expression of CD83 Is Regulated by HuR via a Novel cis-Active Coding Region RNA Element

“…Taking into account the data of the present study, this suggests that the RNA stabilization and nuclear export function of HuR can be separated from each other, depending on the RNA target sequence. According to this, the stabilization of mRNAs depends on the interaction of HuR with an ARE sequence and occurs in the cytoplasm (61,93,94). In contrast, the binding of HuR to the CD83 PRE does not affect the decay of transcripts bearing this RNA sequence.…”

Distinct Domains of AU-Rich Elements Exert Different Functions in mRNA Destabilization and Stabilization by p38 Mitogen-Activated Protein Kinase or HuR

“…Furthermore, suppression of HuR amounts by antisense RNA or small interfering RNA approaches inhibited stabilization induced by hypoxia (23), cytokines (35), or UV light (43) or during cell cycle progression (42). Most recently, constitutively active MK2 (41), as well as stimuli known to activate the p38/MK2 pathway (H 2 O 2 [41] and TNF-␣ [1]), was found to induce cytoplasmic accumulation of HuR. Thus, HuR could play a role in p38/MK2-induced stabilization.…”