Angiotensin II cyclic analogs as tools to investigate AT1R biased signaling mechanisms

Abstract

G protein coupled receptors (GPCRs) produce pleiotropic effects by their capacity to engage numerous signaling pathways once activated. Functional selectivity (also called biased signaling), where specific compounds can bring GPCRs to adopt conformations that enable selective receptor coupling to distinct signaling pathways, continues to be significantly investigated. However, an important but often overlooked aspect of functional selectivity is the capability of ligands such as angiotensin II (AngII) to adopt specific conformations that may preferentially bind to selective GPCRs structures. Understanding both receptor and ligand conformation is of the utmost importance for the design of new drugs targeting GPCRs. In this study, we examined the properties of AngII cyclic analogs to impart biased agonism on the angiotensin type 1 receptor (ATR). Positions 3 and 5 of AngII were substituted for cysteine and homocysteine residues ([SarHcy]AngII, [SarCysHcy]AngII and [SarCys]AngII) and the resulting analogs were evaluated for their capacity to activate the Gq/11, G12, Gi2, Gi3, Gz, ERK and β-arrestin (βarr) signaling pathways via ATR. Interestingly, [SarHcy]AngII exhibited potency and full efficacy on all pathways tested with the exception of the Gq pathway. Molecular dynamic simulations showed that the energy barrier associated with the insertion of residue Phe of AngII within the hydrophobic core of ATR, associated with Gq/11 activation, is increased with [SarHcy]AngII. These results suggest that constraining the movements of molecular determinants within a given ligand by introducing cyclic structures may lead to the generation of novel ligands providing more efficient biased agonism.

Label-free cell signaling pathway deconvolution of angiotensin type 1 receptor reveals time-resolved G-protein activity and distinct AngII and AngIIIIV responses

“…The specific contributions of both G12 and G13 were also assessed in this study using siRNAs. We observed a role for G12 in the early events (phase I) which support recent data by our group using BRET biosensors [39,40] and from other research groups [41]. We also observed a role of G13 in the later phase of the signal (Table 2), where its relative weight in the response observed must be interpreted cautiously given the slight effects of the transfection procedure on the general response profile of AngII.…”

Monitoring TRPC7 Conformational Changes by BRET Following GPCR Activation

“…Since GFP10-TRPC7-RLucII displayed a superior dynamic range upon AT 1 R agonist stimulation (0.057 ± 0.019), which has already been demonstrated to play a role in TRPC7 activation [ 14 ], and as compared to TPα (0.055 ± 0.007) and UT (0.022 ± 0.024), we further characterized this biosensor using AT 1 R, which is known to couple to Gα q , Gα i/o and Gα 12/13 [ 29 , 30 , 31 ]. Therefore, we investigated the effect of pharmacological inhibition of specific AT 1 R pathways by using the Gα q/11 inhibitor YM-254890 (1 µM), the Gα i/o inhibitor pertussis toxin (PTX) (100 ng/mL), and the Rho kinase inhibitor Y27632 (10 µM), a downstream effector of the Gα 12/13 pathway ( Figure 5 a–c).…”

Angiotensin II type 1 receptor localizes at the blood–bile barrier in humans and pigs

“…Alternatively, AT1R can be activated by physiological membrane stretch (Durvasula et al 2004 ; Ramkhelawon et al 2013 ; Mederos y Schnitzler et al 2011 ; De Mello 2012 ). The AT1R interacts with Gα subunits, such as Gαq/11, Gα12/13, and Gαi (St-Pierre et al 2018 ; Shatanawi et al 2011 ). AT1R coupled to Gαq/11 works as cell surface mechanosensor in cardiomyocytes and in smooth muscle cells of small renal and cerebral resistance arteries (Mederos y Schnitzler et al 2011 ).…”