Macrophage activation participates pivotally in the pathophysiology of chronic inflammatory diseases, including atherosclerosis. Through the receptor EP4, prostaglandin E2 (PGE2) 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 PGE2-mediated anti-inflammation remains undefined. Here we demonstrate that PGE2 pretreatment selectively inhibits lipopolysaccharide (LPS)-induced nuclear factor κB1 (NF-κB1) 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-κB 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-κB1 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 PGE2-EP4 signaling on LPS-induced p105 phosphorylation. Additionally, EPRAP knockdown as well as deficiency of NF-κB1 in macrophages attenuates the inhibitory effect of PGE2 on LPS-induced MIP-1β production. Thus, PGE2-EP4 signaling augments NF-κB1 p105 protein stability through EPRAP after proinflammatory stimulation, limiting macrophage activation.