Dosage-dependent regulation of cell prolifration and adhesion through dual beta2-adrenergic receptor/cAMP signals

Abstract

The role of Beta-adrenergic receptors (B-ARs) remains controversial in normal and tumor breast. Herein we explore the cAMP signaling involved in B-AR-dependent control of proliferation and adhesion of nontumor human breast cell line MCF-10A. Low concentrations of a B-agonist, isoproterenol (ISO), promote cell adhesion (87.5% cells remaining adherent to the plastic dishes following specific detachment vs. 35.0% in control, P 0.001), while increasing concentrations further engages an additional 36% inhibition of Erk1/2 phosphorylation (p-Erk1/2) -dependent cell proliferation (P 0.01). Isoproterenol dose response on cell adhesion was fitted to a 2-site curve (EC50(1): 16.5 +/-11.5 fM, EC50(2): 4.08 +/- 3.09 nM), while ISO significantly inhibited p-Erk1/2 according to a 1-site model (EC50: 0.25 +/- 0.13 nM). Using B-AR-selective agonist/antagonists and cAMP analogs/inhibitors, we identified a dosage-dependent signaling in which low ISO concentrations target a B2-AR population localized in raft microdomains and stimulate a Gs/cAMP/Epac/adhesion-signaling module, while higher concentrations engage a concomitant activation of another B2-AR population outside rafts and inhibit the proliferation by a Gs/cAMP/PKA-dependent signaling module. Our data provide a new molecular basis for the dose-dependent switch of B-AR signaling. This study also sheds light on a new cAMP pathway core mechanism with a single receptor triggering dual cAMP signaling controlled by PKA or Epac but with different cellular outputs.