Abstract
For a rapid induction and efficient resolution of the inflammatory response, gene expression in cells of the immune system is tightly regulated at the transcriptional and post-transcriptional level. The control of mRNA translation has emerged as an important determinant of protein levels, yet its role in macrophage activation is not well understood. We systematically analyzed the contribution of translational regulation to the early phase of the macrophage response by polysome fractionation from mouse macrophages stimulated with lipopolysaccharide (LPS). Individual mRNAs whose translation is specifically regulated during macrophage activation were identified by microarray analysis. Stimulation with LPS for 1 h caused translational activation of many feedback inhibitors of the inflammatory response including NF-κB inhibitors (Nfkbid, Nfkbiz, Nr4a1, Ier3), a p38 MAPK antagonist (Dusp1) and post-transcriptional suppressors of cytokine expression (Zfp36 and Zc3h12a). Our analysis showed that their translation is repressed in resting and de-repressed in activated macrophages. Quantification of mRNA levels at a high temporal resolution by RNASeq allowed us to define groups with different expression patterns. Thereby, we were able to distinguish mRNAs whose translation is actively regulated from mRNAs whose polysomal shifts are due to changes in mRNA levels. Active up-regulation of translation was associated with a higher content in AU-rich elements (AREs). For one example, Ier3 mRNA, we show that repression in resting cells as well as de-repression after stimulation depends on the ARE. Bone-marrow derived macrophages from Ier3 knockout mice showed reduced survival upon activation, indicating that IER3 induction protects macrophages from LPS-induced cell death. Taken together, our analysis reveals that translational control during macrophage activation is important for cellular survival as well as the expression of anti-inflammatory feedback inhibitors that promote the resolution of inflammation.
Highlights
In their function as innate immune cells, macrophages are highly sensitive to endogenous and exogenous danger signals
To ensure a rapid but transient inflammatory response, gene expression is regulated at multiple levels, including transcription, stability and translation of mRNAs
While control of transcription and mRNA stability has been studied extensively, little is known about translational regulation in macrophages
Summary
In their function as innate immune cells, macrophages are highly sensitive to endogenous and exogenous danger signals They sense pathogen-associated molecular patterns through Tolllike receptors (TLRs) and mount a tightly controlled immune response. The NF-kB pathway together with the p38 MAPK, ERK1/2 and JNK pathways causes a highly orchestrated, transient induction of numerous inflammatory genes Such dynamic gene expression patterns are achieved by regulation at multiple levels, as exemplified by the pro-inflammatory cytokine TNF. LPS acts at the post-transcriptional level and controls the splicing, nuclear export, stability and translation of Tnf mRNA [2]. In their 39 untranslated region (UTR), many cytokine mRNAs contain an AU-rich element (ARE), which recruits specific RNA-binding proteins [3]. MicroRNAs [3] and a recently discovered stem-loop motif that acts as a constitutive RNA decay element (CDE) [9] further suppress the expression of Tnf and other immune-related mRNAs at the post-transcriptional level
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