RESUMEN
EP2 and EP4 prostanoid receptors have long been considered to have similar roles, since they are known to couple with Gαs-protein and activate cAMP-mediated signaling pathways. In this study, we re-evaluated the results of cAMP assays with or without phosphodiesterase (PDE) inhibitor pretreatment. Here, we show that in the absence of PDE inhibitor pretreatment, prostaglandin E2 causes accumulation of cAMP in EP2 receptors, whereas markedly low levels of cAMP accumulated in EP4 receptors. By applying the Black/Leff operational model calculation, we found that EP2 receptors have a biased ability to intrinsically activate the Gαs-protein-mediated pathway, whereas EP4 receptors have strong biased activity for the Gαi-protein-mediated pathway. Thus, EP2 and EP4 receptors may not be similar Gαs-coupled receptors but instead substantially different receptors with distinct roles.
Asunto(s)
Subtipo EP2 de Receptores de Prostaglandina E , Subtipo EP4 de Receptores de Prostaglandina E , Prostaglandinas , Subtipo EP2 de Receptores de Prostaglandina E/metabolismo , Subtipo EP4 de Receptores de Prostaglandina E/metabolismo , Transducción de Señal/fisiologíaRESUMEN
BACKGROUND: Human DP and EP2 receptors are two of the most homologically related receptors coupling with Gαs-protein, which stimulate adenylyl cyclase to produce cAMP. Indeed, both receptors are considered to be generated by tandem duplication. It has been reported that other highly homologous and closely related ß1- and ß2-adrenergic receptors interact distinctly with and differentially regulate cAMP-specific phosphodiesterase (PDE) 4 recruitment. METHODS: First, we focused on the cAMP degradation pathways of DP and EP2 receptors stimulated by prostaglandin (PG) D2 or PGE2 using HEK cells stably expressing either human DP receptors or EP2 receptors. Then, distances between ligands and amino acids of the receptors were evaluated by molecular dynamics (MD) analysis. RESULTS: We found that PGD2/EP2 receptors exerted a greater effect on PDE4 activity than PGE2/EP2 receptors. Moreover, by MD analysis, either the PGD2 or EP2 receptor was moved and the distance was shortened between them. According to the results, DP receptors retain reactivity for PGE2, but EP2 receptors may be activated only by PGE2, at least in terms of cAMP formation, through the differential functional coupling of PDE4 probably with ß-arrestin. CONCLUSION: Since DP receptors and EP2 receptors are considered to be duplicated genes, DP receptors may still be in a rapid evolutionary stage as a duplicated copy of EP2 receptors and have not yet sufficient selectivity for their cognate ligand, PGD2.
Asunto(s)
Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Dinoprostona/metabolismo , Prostaglandina D2/metabolismo , Subtipo EP2 de Receptores de Prostaglandina E/metabolismo , Adenilil Ciclasas/metabolismo , Línea Celular , AMP Cíclico/metabolismo , Células HEK293 , Humanos , Ligandos , Receptores Inmunológicos/metabolismo , Receptores de Prostaglandina/metabolismo , Transducción de Señal/fisiologíaRESUMEN
Prostaglandin E2 (PGE2) is well-known as an endogenous proinflammatory prostanoid synthesized from arachidonic acid by the activation of cyclooxygenase-2. E type prostanoid (EP) receptors are cognates for PGE2 that have four main subtypes: EP1 to EP4. Of these, the EP2 and EP4 prostanoid receptors have been shown to couple to Gαs-protein and can activate adenylyl cyclase to form cAMP. Studies suggest that EP4 receptors are involved in colorectal homeostasis and cancer development, but further work is needed to identify the roles of EP2 receptors in these functions. After sufficient inflammation has been evoked by PGE2, it is metabolized to 15-keto-PGE2 Thus, 15-keto-PGE2 has long been considered an inactive metabolite of PGE2 However, it may have an additional role as a biased and/or partial agonist capable of taking over the actions of PGE2 to gradually terminate reactions. Here, using cell-based experiments and in silico simulations, we show that PGE2-activated EP4 receptor-mediated signaling may evoke the primary initiating reaction of the cells, which would take over the 15-keto-PGE2-activated EP2 receptor-mediated signaling after PGE2 is metabolized to 15-keto-PGE2 The present results shed light on new aspects of 15-keto-PGE2, which may have important roles in passing on activities to EP2 receptors from PGE2-stimulated EP4 receptors as a "switched agonist." This novel mechanism may be significant for gradually terminating PGE2-evoked inflammation and/or maintaining homeostasis of colorectal tissues/cells functions.