Background and Purpose
Previous studies indicate that prostaglandin EP2 receptors (EP2Rs) selectively couple to adenylyl cyclase type 2 (AC2) in non-lipid raft domains of airway smooth muscle (ASM) cells, where they regulate specific cAMP-dependent responses. The goal of the present study was to identify the cellular microdomains where EP2Rs stimulate cAMP production.
FRET-based cAMP biosensors were targeted to different subcellular locations of primary human ASM cells. The Epac2-camps biosensor, which expresses throughout the cell, was used to measure bulk cytoplasmic responses. Epac2-MyrPalm and Epac2-CAAX were used to measure responses associated with lipid raft and non-raft regions of the plasma membrane, respectively. Epac2-NLS was used to monitor responses at the nucleus.
Activation of AC with forskolin or β2-adrenergic receptors (β2ARs) with isoproterenol increased cAMP in all subcellular locations. Activation of EP2Rs with butaprost produced cAMP responses that were most readily detected by the non-raft and nuclear sensors, but only weakly detected by the cytosolic sensor and not detected at all by the lipid raft sensor. Exposure to rolipram, a phosphodiesterase type 4 (PDE4) inhibitor, unmasked the ability of EP2Rs to increase cAMP levels associated with lipid raft domains. Overexpression of AC2 selectively increased EP2R-stimulated production of cAMP in non-raft membrane domains.
Conclusions and Implications
EP2R activation of AC2 leads to cAMP production in non-raft and nuclear compartments of human ASMs, while β2AR signaling is broadly detected across microdomains. Activity of PDE4 appears to play a role in maintaining the integrity of compartmentalized EP2R responses in these cells.
Agarwal SR, Miyashiro K, Latt H, Ostrom RS, Harvey RD. Compartmentalized cAMP responses to prostaglandin EP2 receptor activation in human airway smooth muscle cells. Br J Pharmacol. 2017. doi: 10.1111/bph.13904