USP20 deletion promotes eccentric cardiac remodeling in response to pressure overload and increases mortality.

Left ventricular hypertrophy (LVH) caused by chronic pressure overload with subsequent pathological remodeling is a major cardiovascular risk factor for heart failure and mortality. The role of deubiquitinases in LVH has not been well characterized. To define whether the deubiquitinase ubiquitin-specific peptidase 20 (USP20) regulates LVH, we subjected USP20 knockout (KO) and cognate wild-type (WT) mice to chronic pressure overload by transverse aortic constriction (TAC) and measured changes in cardiac function by serial echocardiography followed by histological and biochemical evaluations.

Phosphorylation patterns in the AT1R C-terminal tail specify distinct downstream signaling pathways.

Different ligands stabilize specific conformations of the angiotensin II type 1 receptor (AT1R) that direct distinct signaling cascades mediated by heterotrimeric G proteins or β-arrestin. These different active conformations are thought to engage distinct intracellular transducers because of differential phosphorylation patterns in the receptor C-terminal tail (the "barcode" hypothesis). Here, we identified the AT1R barcodes for the endogenous agonist AngII, which stimulates both G protein activation and β-arrestin recruitment, and for a synthetic biased agonist that only stimulates β-arrestin recruitment.

Engineering and Characterization of a Long-Half-Life Relaxin Receptor RXFP1 Agonist.

Relaxin-2 is a peptide hormone with important roles in human cardiovascular and reproductive biology. Its ability to activate cellular responses such as vasodilation, angiogenesis, and anti-inflammatory and antifibrotic effects has led to significant interest in using relaxin-2 as a therapeutic for heart failure and several fibrotic conditions. However, recombinant relaxin-2 has a very short serum half-life, limiting its clinical applications.

G Protein-Coupled Receptors: A Century of Research and Discovery.

GPCRs (G protein-coupled receptors), also known as 7 transmembrane domain receptors, are the largest receptor family in the human genome, with ≈800 members. GPCRs regulate nearly every aspect of human physiology and disease, thus serving as important drug targets in cardiovascular disease. Sharing a conserved structure comprised of 7 transmembrane α-helices, GPCRs couple to heterotrimeric G-proteins, GPCR kinases, and β-arrestins, promoting downstream signaling through second messengers and other intracellular signaling pathways.

G protein-coupled receptors: from radioligand binding to cellular signaling.

Radioligand binding techniques facilitated the identification and study of G-protein coupled receptors that now represent the largest class of targets for therapeutic drugs.

Pathophysiology and pharmacology of G protein-coupled receptors in the heart.

G protein-coupled receptors (GPCRs), comprising the largest superfamily of cell surface receptors, serve as fundamental modulators of cardiac health and disease owing to their key roles in the regulation of heart rate, contractile dynamics, and cardiac function. Accordingly, GPCRs are heavily pursued as drug targets for a wide variety of cardiovascular diseases ranging from heart failure, cardiomyopathy, and arrhythmia to hypertension and coronary artery disease. Recent advancements in understanding the signalling mechanisms, regulation, and pharmacological properties of GPCRs have provided valuable insights that will guide the development of novel therapeutics.

G protein-coupled receptor signaling: transducers and effectors.

G protein-coupled receptors (GPCRs) are of considerable interest due to their importance in a wide range of physiological functions and in a large number of Food and Drug Administration (FDA)-approved drugs as therapeutic entities. With continued study of their function and mechanism of action, there is a greater understanding of how effector molecules interact with a receptor to initiate downstream effector signaling. This review aims to explore the signaling pathways, dynamic structures, and physiological relevance in the cardiovascular system of the three most important GPCR signaling effectors: heterotrimeric G proteins, GPCR kinases (GRKs), and β-arrestins.

Proximity labeling for investigating protein-protein interactions.

The study of protein complexes and protein-protein interactions is of great importance due to their fundamental roles in cellular function. Proximity labeling, often coupled with mass spectrometry, has become a powerful and versatile tool for studying protein-protein interactions by enriching and identifying proteins in the vicinity of a specified protein-of-interest. Here, we describe and compare traditional approaches to investigate protein-protein interactions to current day state-of-the-art proximity labeling methods.

Late onset cardiovascular dysfunction in adult mice resulting from galactic cosmic ray exposure.

The complex and inaccessible space radiation environment poses an unresolved risk to astronaut cardiovascular health during long-term space exploration missions. To model this risk, healthy male c57BL/6 mice aged six months (corresponding to an astronaut of 34 years) were exposed to simplified galactic cosmic ray (GCR; 5-ion sim) irradiation at the NASA Space Radiation Laboratory (NSRL) at Brookhaven National Laboratories (BNL). Multi-modal cardiovascular functional assessments performed longitudinally and terminally revealed significant impairment in cardiac function in mice exposed to GCR compared to unirradiated controls, gamma irradiation, or single mono-energetic ions (Fe or O).

-Arrestin-Biased Allosteric Modulator Potentiates Carvedilol-Stimulated Adrenergic Receptor Cardioprotection.

adrenergic receptors ( ARs) are central regulators of cardiac function and a drug target for cardiac disease. As a member of the G protein-coupled receptor family, ARs activate cellular signaling by primarily coupling to Gs proteins to activate adenylyl cyclase, cAMP-dependent pathways, and the multifunctional adaptor-transducer protein -arrestin. Carvedilol, a traditional -blocker widely used in treating high blood pressure and heart failure by blocking adrenergic receptor-mediated G protein activation, can selectively stimulate Gs-independent -arrestin signaling of adrenergic receptors, a process known as -arrestin-biased agonism.

Mapping Angiotensin II Type 1 Receptor-Biased Signaling Using Proximity Labeling and Proteomics Identifies Diverse Actions of Biased Agonists.

Angiotensin II type 1 receptors (AT1Rs) are one of the most widely studied G-protein-coupled receptors. To fully appreciate the diversity in cellular signaling profiles activated by AT1R transducer-biased ligands, we utilized peroxidase-catalyzed proximity labeling to capture proteins in close proximity to AT1Rs in response to six different ligands: angiotensin II (full agonist), S1I8 (partial agonist), TRV055 and TRV056 (G-protein-biased agonists), and TRV026 and TRV027 (β-arrestin-biased agonists) at 90 s, 10 min, and 60 min after stimulation (ProteomeXchange Identifier PXD023814). We systematically analyzed the kinetics of AT1R trafficking and determined that distinct ligands lead AT1R to different cellular compartments for downstream signaling activation and receptor degradation/recycling.

Synthetic nanobodies as angiotensin receptor blockers.

There is considerable interest in developing antibodies as functional modulators of G protein-coupled receptor (GPCR) signaling for both therapeutic and research applications. However, there are few antibody ligands targeting GPCRs outside of the chemokine receptor group. GPCRs are challenging targets for conventional antibody discovery methods, as many are highly conserved across species, are biochemically unstable upon purification, and possess deeply buried ligand-binding sites.

The β-arrestin-biased β-adrenergic receptor blocker carvedilol enhances skeletal muscle contractility.

A decrease in skeletal muscle strength and functional exercise capacity due to aging, frailty, and muscle wasting poses major unmet clinical needs. These conditions are associated with numerous adverse clinical outcomes including falls, fractures, and increased hospitalization. Clenbuterol, a β-adrenergic receptor (βAR) agonist enhances skeletal muscle strength and hypertrophy; however, its clinical utility is limited by side effects such as cardiac arrhythmias mediated by G protein signaling.

A murine model of increased coronary sinus pressure induces myocardial edema with cardiac lymphatic dilation and fibrosis.

Myocardial edema is a consequence of many cardiovascular stressors, including myocardial infarction, cardiac bypass surgery, and hypertension. The aim of this study was to establish a murine model of myocardial edema and elucidate the response of cardiac lymphatics and the myocardium. Myocardial edema without infarction was induced in mice by cauterizing the coronary sinus, increasing pressure in the coronary venous system, and inducing myocardial edema.

β-arrestin 1 regulates β2-adrenergic receptor-mediated skeletal muscle hypertrophy and contractility.

Background: β-adrenergic receptors (βARs) are the target of catecholamines and play fundamental roles in cardiovascular, pulmonary, and skeletal muscle physiology. An important action of βAR stimulation on skeletal muscle is anabolic growth, which has led to the use of agonists such as clenbuterol by athletes to enhance muscle performance. While previous work has demonstrated that βARs can engage distinct signaling and functional cascades mediated by either G proteins or the multifunctional adaptor protein, β-arrestin, the precise role of β-arrestin in skeletal muscle physiology is not known.

The deubiquitinase ubiquitin-specific protease 20 is a positive modulator of myocardial β-adrenergic receptor expression and signaling.

Reversible ubiquitination of G protein-coupled receptors regulates their trafficking and signaling; whether deubiquitinases regulate myocardial β-adrenergic receptors (βARs) is unknown. We report that ubiquitin-specific protease 20 (USP20) deubiquitinates and attenuates lysosomal trafficking of the βAR. βAR-induced phosphorylation of USP20 Ser-333 by protein kinase A-α (PKAα) was required for optimal USP20-mediated regulation of βAR lysosomal trafficking.

G-Protein-Coupled Receptors in Heart Disease.

GPCRs (G-protein [guanine nucleotide-binding protein]-coupled receptors) play a central physiological role in the regulation of cardiac function in both health and disease and thus represent one of the largest class of surface receptors targeted by drugs. Several antagonists of GPCRs, such as βARs (β-adrenergic receptors) and Ang II (angiotensin II) receptors, are now considered standard of therapy for a wide range of cardiovascular disease, such as hypertension, coronary artery disease, and heart failure. Although the mechanism of action for GPCRs was thought to be largely worked out in the 80s and 90s, recent discoveries have brought to the fore new and previously unappreciated mechanisms for GPCR activation and subsequent downstream signaling.

Manifold roles of β-arrestins in GPCR signaling elucidated with siRNA and CRISPR/Cas9.

G protein-coupled receptors (GPCRs) use diverse mechanisms to regulate the mitogen-activated protein kinases ERK1/2. β-Arrestins (βArr1/2) are ubiquitous inhibitors of G protein signaling, promoting GPCR desensitization and internalization and serving as scaffolds for ERK1/2 activation. Studies using CRISPR/Cas9 to delete βArr1/2 and G proteins have cast doubt on the role of β-arrestins in activating specific pools of ERK1/2.

The two-pore domain potassium channel TREK-1 mediates cardiac fibrosis and diastolic dysfunction.

Cardiac two-pore domain potassium channels (K2P) exist in organisms from Drosophila to humans; however, their role in cardiac function is not known. We identified a K2P gene, CG8713 (sandman), in a Drosophila genetic screen and show that sandman is critical to cardiac function. Mice lacking an ortholog of sandman, TWIK-related potassium channel (TREK-1, also known Kcnk2), exhibit exaggerated pressure overload-induced concentric hypertrophy and alterations in fetal gene expression, yet retain preserved systolic and diastolic cardiac function.