Nuclear RSK1 Induces Pro-inflammatory Activation of Macrophages through STAT1 Phosphorylation at Ser727

Objective: Maladaptive inflammatory responses involve macrophage activation by pro-inflammatory cytokines such as IFN-γ. Proteins that undergo nuclear translocation may regulate these processes. Approach and Results: To explore novel key regulators of macrophage activation, we performed quantitative proteomics to monitor protein translocation to the nuclei of human primary macrophages elicited with IFN-γ for 0, 10, 20, 30, or 60 min. Bioinformatics identified RSK1, a ribosomal protein kinase, as one of the candidates. We found that IFN-γ stimulation promotes RSK1 phosphorylation at Ser380. STAT1 is a key mediator of IFN-γ-triggered cellular responses. Mass spectrometry identified RSK1-mediated phosphorylation sites within STAT1, including Ser727. siRNA silencing of RSK1 attenuated STAT1 phosphorylation in IFN-γ-stimulated macrophages. In concert with these results, RSK1 silencing hindered IFN-γ-induced secretion of pro-inflammatory chemokines in human primary macrophages, such as CCL2/MCP-1, CCL7/MCP-3, and CCL8/MCP-2. Conclusion: We discovered that RSK1 nuclear translocation triggers STAT1 phosphorylation, resulting in pro-inflammatory activation of macrophages (Figure), a novel role for a ribosomal protein-associated kinase in nuclear signaling and inflammation.