Deciphering the Novel Role of Gαz in Mu Opioid Receptor Signaling

Abstract

A better understanding of mu opioid receptor (MOR) signaling is necessary to combat the negative side effects of opioid analgesics. The MOR is a G protein coupled receptor that couples to the Gαi/o class of proteins; however, it is common to downplay the diversity within this class. Gαz is an often-overlooked member of this class, yet its prevalent expression in similar regions of the brain as opioid receptors suggests it may serve a distinct role in MOR signaling. Using a bioluminescence resonance energy transfer (BRET) sensor, we investigated the potential of Gαz and its regulator of G protein signaling counterpart, RGSZ, as a means to control both efficacy and potency of MOR agonists. To avoid hindering the MOR•Gαz interaction, Gα signaling was correlated to the interaction of Venus-tagged Gβγ with a non-functional, C-terminal fragment of its effector, G protein-coupled receptor kinase 3 (GRK3ct). As a result, agonist binding to MOR resulted in an increase in BRET due to G-protein dissociation. The full agonists DAMGO, β-endorphin, methadone, and fentanyl were more potent when MOR signaled through Gαz compared to other Gαi/o subunits tested. For example, the EC50 value was 3-fold greater when the MOR signaled through Gαi1 compared to Gαz. Interestingly, fentanyl was equally efficacious with similar Emax values with both Gα subunits. Additionally, partial agonists such as morphine, (-)pentazocine, and (-)cyclazocine were both more potent and more efficacious when the MOR signaled through Gαz compared to the other Gαi/o subunits. In the case of (-)pentazocine, when MOR signaled through Gαz, the EC50 value was 2-fold lower compared to when MOR signaled through Gαi1. Moreover, the Emax value was also 2-fold higher when the MOR signaled through Gαz compared to Gαi1. Next, RGSZ was transfected and again the BRET assay was used to determine potency and efficacy. When RGSZ was present in the system, the concentration-response curve of DAMGO had a significant rightward shift. Taken together, Gαz can enhance opioid potency and efficacy downstream of receptor binding.