Showing papers by "Robert J. Lefkowitz published in 2021"
TL;DR: In this paper, the authors have solved the structure of a GPCR-G protein-βarr megacomplex by cryo-electron microscopy, which illustrates the independent and simultaneous coupling of G protein to the receptor intracellular core, and binding of a βarr to a phosphorylated receptor C-terminal tail.
Abstract: G protein-coupled receptors (GPCRs) are a large class of cell-surface receptor involved in cellular signaling that are currently the target of over one third of all clinically approved therapeutics. Classically, an agonist-bound, active GPCR couples to and activates G proteins through the receptor intracellular core. To attenuate G protein signaling, the GPCR is phosphorylated at its C-terminal tail and/or relevant intracellular loops, allowing for the recruitment of β-arrestins (βarrs). βarrs then couple to the receptor intracellular core in order to mediate receptor desensitization and internalization. However, our laboratory and others have observed that some GPCRs are capable of continuously signaling through G protein even after internalization. This mode of sustained signaling stands in contrast with our previous understanding of GPCR signaling, and its molecular mechanism is still not well understood. Recently, we have solved the structure of a GPCR-G protein-βarr megacomplex by cryo-electron microscopy. This 'megaplex' structure illustrates the independent and simultaneous coupling of a G protein to the receptor intracellular core, and binding of a βarr to a phosphorylated receptor C-terminal tail, with all three components maintaining their respective canonically active conformations. The structure provides evidence for the ability of a GPCR to activate G protein even while being bound to and internalized by βarr. It also reveals that the binding of G protein and βarr to the same GPCR is not mutually exclusive, and raises a number of future questions to be answered regarding the mechanism of sustained signaling.
17 citations
TL;DR: In this article, a small molecule library screen against β2 adrenergic receptors (β2AR) identified Compound-6 (Cmpd-6) to be a positive allosteric modulator for β2ARs.
Abstract: β1 adrenergic receptors (β1ARs) are central regulators of cardiac function and a drug target for cardiac disease. As a member of G protein-coupled receptor family, β1ARs activate cellular signaling by primarily coupling to Gs proteins to activate adenylyl cyclase and cAMP-dependent pathways, and the multifunctional adaptor-transducer protein β-arrestin. Carvedilol, a traditional β-blocker widely used in treating high blood pressure and heart failure by blocking βAR-mediated G-protein activation, can selectively stimulate Gs-independent β-arrestin signaling of βARs, a process known as β-arrestin-biased agonism. Recently a DNA-encoded small molecule library screen against agonist-occupied β2 adrenergic receptors (β2AR) identified Compound-6 (Cmpd-6) to be a positive allosteric modulator for agonists on β2ARs. Intriguingly, it was further discovered that Cmpd-6 is positively cooperative with the β-arrestin biased ligand carvedilol at β2ARs. Here we describe the surprising finding that at β1ARs, unlike the case of β2ARs, Cmpd-6 is cooperative only with carvedilol and not agonists. Cmpd-6 increases the binding affinity of carvedilol for β1ARs and potentiates carvedilol-stimulated, β-arrestin-dependent β1AR signaling such as epidermal growth factor receptor transactivation and extracellular signal-regulated kinase activation, while having no effect on Gs-mediated cAMP generation. In vivo, Cmpd-6 enhances the anti-apoptotic cardioprotective effect of carvedilol in response to myocardial ischemia/reperfusion injury. This anti-apoptotic role of carvedilol is dependent on β-arrestins, since it is lost in mice with myocyte-specific deletion of β-arrestins. Our findings demonstrate that Cmpd-6 is a selective β-arrestin-biased allosteric modulator of β1ARs and highlight its potential clinical utility in enhancing carvedilol-mediated cardioprotection against ischemic injury. Significance Statement In this study, we demonstrate the positive cooperativity of Cmpd-6 on β1ARs as a β-arrestin-biased positive allosteric modulator. Cmpd-6 selectively enhances the affinity and cellular signaling of carvedilol, a known β-arrestin-biased β-blocker for β1ARs, while having minimal effect on other ligands tested. Importantly, Cmpd-6 enhances the β-arrestin-dependent in vivo cardioprotective effect of carvedilol during ischemia/reperfusion injury-induced apoptosis. Our data support the potential therapeutic application of Cmpd-6 to enhance the clinical benefits of carvedilol in the treatment of cardiac disease.
14 citations
TL;DR: In this article, a small molecule-mediated allosteric modulation of the β-arrestin-biased β-blocker, carvedilol, was reported, which has potential implications in advancing G-protein-coupled receptor allostery, developing biased therapeutics and remedying cardiovascular ailments.
Abstract: Among β-blockers that are clinically prescribed for heart failure, carvedilol is a first-choice agent with unique pharmacological properties. Carvedilol is distinct from other β-blockers in its ability to elicit β-arrestin-biased agonism, which has been suggested to underlie its cardioprotective effects. Augmenting the pharmacologic properties of carvedilol thus holds the promise of developing more efficacious and/or biased β-blockers. We recently identified compound-6 (cmpd-6), the first small molecule positive allosteric modulator of the β2-adrenergic receptor (β2AR). Cmpd-6 is positively cooperative with orthosteric agonists at the β2AR and enhances agonist-mediated transducer (G-protein and β-arrestin) signaling in an unbiased manner. Here, we report that cmpd-6, quite unexpectedly, displays strong positive cooperativity only with carvedilol among a panel of structurally diverse β-blockers. Cmpd-6 enhances the binding affinity of carvedilol for the β2AR and augments its ability to competitively antagonize agonist-induced cAMP generation. Cmpd-6 potentiates β-arrestin1- but not Gs-protein-mediated high-affinity binding of carvedilol at the β2AR and β-arrestin-mediated cellular functions in response to carvedilol including extracellular signal-regulated kinase phosphorylation, receptor endocytosis, and trafficking into lysosomes. Importantly, an analog of cmpd-6 that selectively retains positive cooperativity with carvedilol acts as a negative modulator of agonist-stimulated β2AR signaling. These unprecedented cooperative properties of carvedilol and cmpd-6 have implications for fundamental understanding of G-protein-coupled receptor (GPCR) allosteric modulation, as well as for the development of more effective biased beta blockers and other GPCR therapeutics. SIGNIFICANCE STATEMENT: This study reports on the small molecule-mediated allosteric modulation of the β-arrestin-biased β-blocker, carvedilol. The small molecule, compound-6 (cmpd-6), displays an exclusive positive cooperativity with carvedilol among other β-blockers and enhances the binding affinity of carvedilol for the β2-adrenergic receptor. Cooperative effects of cmpd-6 augment the β-blockade property of carvedilol while potentiating its β-arrestin-mediated signaling functions. These findings have potential implications in advancing G-protein-coupled receptor allostery, developing biased therapeutics and remedying cardiovascular ailments.
13 citations
TL;DR: In this paper, the authors demonstrate the direct activation of C-Raf by GPCR-βarr1 complexes in vitro and show that βarr1 in complex with a synthetic phosphopeptide mimicking the human V2 vasopressin receptor tail that binds and functionally activates βarrs also allosterically activates the mitogen-activated protein kinase pathway.
2 citations