Allosteric interactions between the oxytocin receptor and the β _2-adrenergic receptor in the modulation of ERK1/2 activation are mediated by heterodimerization

摘要

The oxytocin receptor (OTR) and the β _2-adrenergic receptor (β _2AR) are key regulators of uterine contraction. These two receptors are targets of tocolytic agents used to inhibit pre-term labor. Our recent study on the nature of OTR- and β _2AR-mediated ERK1/2 activation in human hTERT-C3 myometrial cells suggested the presence of an OTR/β _2AR hetero-oligomeric complex (see companion article). The goal of this study was to investigate potential allosteric interactions between OTR and β _2AR and establish the nature of the interactions between these receptors in myometrial cells. We found that OTR-mediated ERK1/2 activation was attenuated significantly when cells were pretreated with the β _2AR agonist isoproterenol or two antagonists, propranolol or timolol. In contrast, pretreatment of cells with a third β _2AR antagonist, atenolol resulted in an increase in OTR-mediated ERK1/2 activation. Similarly, β 2AR-mediated ERK1/2 activation was strongly attenuated by pretreatment with the OTR antagonists, atosiban and OTA. Physical interactions between OTR and β _2AR were demonstrated using co-immunoprecipitation, bioluminescence resonance energy transfer (BRET) and protein-fragment complementation (PCA) assays in HEK 293 cells, the latter experiments indicating the interactions between the two receptors were direct. Our analyses suggest physical interactions between OTR and β _2AR in the context of a new heterodimer pair lie at the heart of the allosteric effects.