Epac1-mediated Rap1 activation is not required for the production of nitric oxide in BV2, murine microglial cells.

This study demonstrates that cyclic AMP (cAMP) production is induced by lipopolysaccharide (LPS) stimulation and activates two different pathways in murine BV2 microglial cells. Two principal effector proteins for cAMP are protein kinase A (PKA) and cAMP-responsive guanine nucleotide exchange factor (Epac), a Rap GDP exchange factor. When cells were treated with various cAMP level modulators, nitric oxide (NO) production increased as the result of posttreatment with Type IV phosphodiesterase (PDE4) inhibitor, rolipram or dibutyryl-cAMP (dbcAMP), at 2 hr after LPS stimulation. Intracellular cAMP increased due to LPS stimulation and the cAMP modulators phosphorylate transcription factor CREB, which is enhanced in turn by posttreatment with dbcAMP. In contrast, the Epac-specific cAMP analog 8-(4-chloro-phenylthio)-2'-O-methyladenosine-3',5'-cyclic monophosphate (8CPT-2Me-cAMP) activates Rap1 in the BV2 cells, but does not induce PKA activation, as judged by CREB phosphorylation. NO production was enhanced by posttreatment with dbcAMP but not by treatment with 8CPT-2Me-cAMP. This suggests that LPS-stimulated NO production is mainly PKA-dependent and also that Epac1-mediated Rap1 activation is not required for the induction of NO production.

Moon EY ，Oh SY ，Han GH ，Lee CS ，Park SK ... -  《JOURNAL OF NEUROSCIENCE RESEARCH》

Lipopolysaccharide stimulates Epac1-mediated Rap1/NF-kappaB pathway in Raw 264.7 murine macrophages.

Moon EY ，Pyo S 《IMMUNOLOGY LETTERS》

ROS/Epac1-mediated Rap1/NF-kappaB activation is required for the expression of BAFF in Raw264.7 murine macrophages.

B-cell activating factor (BAFF) plays a role for the maturation and the maintenance of B cells. Lipopolysaccharide (LPS) activates toll-like receptor 4 (TLR4)-dependent signal transduction, which resulted in BAFF expression through nuclear factor kappa B (NF-κB) activation. Here, we investigated whether BAFF expression could be regulated by p65 phosphorylation through the production of reactive oxygen species (ROS) or cyclic AMP (cAMP) in Raw264.7 murine macrophages. mBAFF expression was reduced by ROS scavengers and it was increased by dibutyl-cAMP, a cAMP analogue. mBAFF expression and mBAFF promoter activity were increased by co-transfection of p65 but they were reduced by p65-small interference (si) RNA. Serine (Ser) 276 phosphorylation of p65 was increased by LPS-mediated PKA activation or by the treatment with forskolin, adenylate cyclase activator and dibutyl-cAMP. In contrast, p65 phosphorylation at Ser276 was decreased by ROS scavengers. H(2)O(2) increased intracellular cAMP concentration, significantly. While no increase in p65 phosphorylation at Ser276 was detected by the treatment with H(2)O(2), CREB and p65 phosphorylation at Ser133 and Ser536 was observed, respectively. It implicates that p65 phosphorylation at Ser276 is independent of ROS-induced cAMP production. As another cAMP effector protein was cAMP-responsive guanine nucleotide exchange factor (Epac), a Rap GDP exchange factor, NF-κB was activated by the treatment with 8-(4-chloro-phenylthio)-2'-O-methyladenosine-3',5'-cyclic monophosphate (CPT) that is an activator to Epac. Epac1-mediated Rap1 was activated by the treatment with H(2)O(2) but it was inhibited by ROS scavengers. CPT induced p65 phosphorylation at both Ser276 and Ser536. CPT also increased not only mBAFF expression but mBAFF promoter activity. Data demonstrate that TLR4-mediated mBAFF expression was resulted from the crosstalk of p65 phosphorylation at Ser536 and Ser276 through ROS- and/or cAMP-mediated signal transduction. It suggests for the first time that ROS/Epac1-mediated Rap1/NF-κB pathway could be required for BAFF expression.

Moon EY ，Lee JH ，Lee JW ，Song JH ，Pyo S ... -  《-》

Cyclic AMP induces integrin-mediated cell adhesion through Epac and Rap1 upon stimulation of the beta 2-adrenergic receptor.

Rangarajan S ，Enserink JM ，Kuiperij HB ，de Rooij J ，Price LS ，Schwede F ，Bos JL ... -  《-》

Cyclic AMP-dependent, protein kinase A-independent activation of extracellular signal-regulated kinase 1/2 following adenosine receptor stimulation in human umbilical vein endothelial cells: role of exchange protein activated by cAMP 1 (Epac1).

Fang Y ，Olah ME 《JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS》