Prostaglandin e receptor type 4-associated protein interacts directly with NF-κB1 and attenuates macrophage activation

Macrophage activation participates pivotally in the pathophysiology of chronic inflammatory diseases, including atherosclerosis. Through the receptor EP4, prostaglandin E-2 (PGE(2)) exerts an anti-inflammatory action in macrophages, suppressing stimulus-induced expression of certain proinflammatory genes, including chemokines. We recently identified a novel EP4 receptor-associated protein (EPRAP), whose function in PGE(2)-mediated anti-inflammation remains undefined. Here we demonstrate that PGE(2) pretreatment selectively inhibits lipopolysaccharide (LPS)-induced nuclear factor kappa B1 (NF-kappa B1) p105 phosphorylation and degradation in mouse bone marrow-derived macrophages through EP4-dependent mechanisms. Similarly, directed EPRAP expression in RAW264.7 cells suppresses LPS-induced p105 phosphorylation and degradation, and subsequent activation of mitogen-activated protein kinase kinase 1/2. Forced expression of EPRAP also inhibits NF-kappa B activation induced by various proinflammatory stimuli in a concentration-dependent manner. In co-transfected cells, EPRAP, which contains multiple ankyrin repeat motifs, directly interacts with NF-kappa B1 p105/p50 and forms a complex with EP4. In EP4-overexpressing cells, PGE2 enhances the protective action of EPRAP against stimulus-induced p105 phosphorylation, whereas EPRAP silencing in RAW264.7 cells impairs the inhibitory effect of PGE(2)-EP4 signaling on LPS-induced p105 phosphorylation. Additionally, EPRAP knockdown as well as deficiency of NF-kappa B1 in macrophages attenuates the inhibitory effect of PGE2 on LPS-induced MIP-1 beta production. Thus, PGE2-EP4 signaling augments NF-kappa B1 p105 protein stability through EPRAP after proinflammatory stimulation, limiting macrophage activation.