Spontaneous Human B2 Bradykinin Receptor Activity Determines the Action of Partial Agonists as Agonists or Inverse Agonists

Abstract

In this report, we show that desensitization regulates ligand-independent, spontaneous activity of the human B2 bradykinin (BK) receptor, and the level of spontaneous receptor activity determines the action of the BK antagonists and partial receptor agonists NPC17731 and HOE140 as agonists or inverse agonists. Spontaneous receptor activity was monitored by measuring basal cellular phosphoinositide (PI) hydrolysis as a function of the density of the receptor in transiently transfected HEK293 cells. Minimal spontaneous activity of the wildtype B2 receptor was detected in these cells. Mutating a cluster of serines and threonines within the fourth intracellular domain of the receptor, which is critical for agonist-promoted desensitization, significantly increased the spontaneous receptor activity. BK, the natural B2 receptor ligand and, consequently, a full agonist, stimulated PI hydrolysis at high and low levels of spontaneous receptor activity. On the other hand, the partial agonists NPC17731 and HOE140 were stimulatory, or agonists, at the lower level of receptor activity but inhibitory, or inverse agonists, at the higher level of activity. These results show that receptors are desensitized in response to their spontaneous activity. Furthermore, these results, which refute traditional theories, show that the capacity of a drug to modulate a receptor response is not intrinsic to the drug but is also dependent on the cellular environment in which the drug acts.GPCR 1 constitute the largest family of receptors and mediate responses to numerous agonists including hormones, neurotransmitters, and sensory stimuli. Receptor activation by the agonist ligand is considered the first step in receptor signal transduction. However, the recent discovery that GPCR exhibit ligand-independent, spontaneous, and mutation-induced activity has introduced a new function that needs to be considered in cellular signaling and ligand action (1) and that can lead to disease (2-4).GPCR have evolved as the most common targets for therapeutic drugs (5). Drug efficacy is defined as the capacity of a drug to activate or inactivate a receptor response and is fundamental to pharmacology and drug development. Classically, this parameter has been considered an intrinsic property of the drug and independent of the cellular environment in which the drug acts (6 -8). Partial agonists, or mixed agonists/antagonists, are prevalent among GPCR drugs and are defined as ligands that elicit a submaximal receptor response and block the response to the natural receptor ligand, a full agonist, or any other ligand of higher efficacy. It has been predicted that in the absence of a more efficacious agonist, the efficacy and behavior of a partial agonist should be directly dependent upon the level of spontaneous receptor activity (9). In other words, if the partial agonist ligand elicits a response that is higher than the spontaneous activity, then the ligand should behave as an agonist, whereas if the ligand elicits a lower response, then the ligand should block t...

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G protein-coupled receptors sense fluid shear stress in endothelial cells

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Spontaneous Formation of a Proteolytic B1 and B2 Bradykinin Receptor Complex with Enhanced Signaling Capacity

“…This response remained B1R-specific because desArg 10 [Leu 9 ]kallidin, a highly selective B1 antagonist, effectively inhibited the response, whereas the selective B2 inverse agonist NPC17731 did not inhibit the response (data not shown). Furthermore, NPC17731 had no effect on the increase in basal PI hydrolysis caused by coexpression, excluding the conversion of B2R to a more active conformational state as the source of this increase (20,21). These results show that the spontaneous interaction of B1R and B2R is functionally significant and translates into an increased coupling efficacy of the receptor to G q and subsequent phospholipase C␤ activity.…”