Both knock‐down and overexpression of Rap2a small GTPase in macrophages result in impairment of NF‐&kgr;B activity and inflammatory gene expression

Abstract

&NA; Small Ras GTPases are key molecules that regulate a variety of cellular responses in different cell types. Rap1 plays important functions in the regulation of macrophage biology during inflammation triggered by toll‐like receptors (TLRs). However, despite sharing a relatively high degree of similarity with Rap1, no studies concerning Rap2 in macrophages and innate immunity have been reported yet. In this work, we show that either way alterations in the levels of Rap2a hampers proper macrophages response to TLR stimulation. Rap2a is activated by LPS in macrophages, and although putative activator TLR‐inducible Ras guanine exchange factor RasGEF1b was sufficient to induce, it was not fully required for Rap2a activation. Silencing of Rap2a impaired LPS‐induced production of IL‐6 cytokine and KC/Cxcl1 chemokine, and also NF‐&kgr;B activity as measured by reporter gene studies. Surprisingly, overexpression of Rap2a did also lead to marked inhibition of NF‐&kgr;B activation induced by LPS, Pam3CSK4 and downstream TLR signaling molecules. We also found that Rap2a can inhibit the LPS‐induced phosphorylation of the NF‐&kgr;B subunit p65 at serine 536. Collectively, our data suggest that expression levels of Rap2a in macrophages might be tightly regulated to avoid unbalanced immune response. Our results implicate Rap2a in TLR‐mediated responses by contributing to balanced NF‐&kgr;B activity status in macrophages.