A neurodevelopmental disorder mutation locks G proteins in the transitory pre-activated state.

Many neurotransmitter receptors activate G proteins through exchange of GDP for GTP. The intermediate nucleotide-free state has eluded characterization, due largely to its inherent instability. Here we characterize a G protein variant associated with a rare neurological disorder in humans.

Internalized β2-Adrenergic Receptors Oppose PLC-Dependent Hypertrophic Signaling.

Background: Chronically elevated neurohumoral drive, and particularly elevated adrenergic tone leading to β-adrenergic receptor (β-AR) overstimulation in cardiac myocytes, is a key mechanism involved in the progression of heart failure. β1-AR (β1-adrenergic receptor) and β2-ARs (β2-adrenergic receptor) are the 2 major subtypes of β-ARs present in the human heart; however, they elicit different or even opposite effects on cardiac function and hypertrophy. For example, chronic activation of β1-ARs drives detrimental cardiac remodeling while β2-AR signaling is protective.

Biasing G Downstream Signaling with Gallein Inhibits Development of Morphine Tolerance and Potentiates Morphine-Induced Nociception in a Tolerant State.

Opioid analgesics are widely used as a treatment option for pain management and relief. However, the misuse of opioid analgesics has contributed to the current opioid epidemic in the United States. Prescribed opioids such as morphine, codeine, oxycodone, and fentanyl are mu-opioid receptor (MOR) agonists primarily used in the clinic to treat pain or during medical procedures, but development of tolerance limits their utility for treatment of chronic pain.

Functional interrogation of cellular Lp(a) uptake by genome-scale CRISPR screening.

An elevated level of lipoprotein(a), or Lp(a), in the bloodstream has been causally linked to the development of atherosclerotic cardiovascular disease and calcific aortic valve stenosis. Steady state levels of circulating lipoproteins are modulated by their rate of clearance, but the identity of the Lp(a) uptake receptor(s) has been controversial. In this study, we performed a genome-scale CRISPR screen to functionally interrogate all potential Lp(a) uptake regulators in HuH7 cells.

A PACAP-activated network for secretion requires coordination of Ca influx and Ca mobilization.

Chromaffin cells of the adrenal medulla transduce sympathetic nerve activity into stress hormone secretion. The two neurotransmitters principally responsible for coupling cell stimulation to secretion are acetylcholine and pituitary adenylate activating polypeptide (PACAP). In contrast to acetylcholine, PACAP evokes a persistent secretory response from chromaffin cells.

Towards an extensive set of criteria for safety and cyber-security evaluation of cyber-physical systems.

Verification and validation (V&V) are complex processes combining different approaches and incorporating many different methods including many activities. System engineers regularly face the question if their V&V activities lead to better products, and having appropriate criteria at hand for evaluation of safety and cybersecurity of the systems would help to answer such a question. Additionally, when there is a demand to improve the quality of an already managed V&V process, there is a struggle over what criteria to use in order to measure the improvement.

Biophysical characterization of the CXC chemokine receptor 2 ligands.

The chemokines of the immune system act as first responders by operating as chemoattractants, directing immune cells to specific locations of inflamed tissues. This promiscuous network is comprised of 50 ligands and 18 receptors where the ligands may interact with the receptors in various oligomeric states i.e.

Stabilization of interdomain interactions in G protein α subunits as a determinant of Gα subtype signaling specificity.

Highly homologous members of the Gα family, Gα, have distinct tissue distributions and physiological functions, yet their biochemical and functional properties are very similar. We recently identified PDZ-RhoGEF (PRG) as a novel Gα effector that is poorly activated by Gα. In a proteomic proximity labeling screen we observed a strong preference for Gα relative to Gα with respect to engagement of a broad range of potential targets.

Phospholipase C epsilon-1 (PLCƐ1) mediates macrophage activation and protection against tuberculosis.

Tuberculosis (TB) caused by () infects up to a quarter of the world's population. Although immune responses can control infection, 5%-10% of infected individuals can progress to active TB disease (progressors). A myriad of host factors regulate disease progression in TB and a better understanding of immune correlates of protection and disease is pivotal for the development of new therapeutics.

A PACAP-activated network for secretion requires coordination of Ca influx and Ca mobilization.

Chromaffin cells of the adrenal medulla transduce sympathetic nerve activity into stress hormone secretion. The two neurotransmitters principally responsible for coupling cell stimulation to secretion are acetylcholine and pituitary adenylate activating polypeptide (PACAP). In contrast to acetylcholine, PACAP evokes a persistent secretory response from chromaffin cells.

Internalized β2-Adrenergic Receptors Inhibit Subcellular Phospholipase C-Dependent Cardiac Hypertrophic Signaling.

Chronically elevated neurohumoral drive, and particularly elevated adrenergic tone leading to β-adrenergic receptor (β-AR) overstimulation in cardiac myocytes, is a key mechanism involved in the progression of heart failure. β1-AR and β2-ARs are the two major subtypes of β-ARs present in the human heart, however, they elicit different or even opposite effects on cardiac function and hypertrophy. For example, chronic activation of β1ARs drives detrimental cardiac remodeling while β2AR signaling is protective.

Stabilization of Interdomain Interactions in G protein α Subunits Determines Gα Subtype Signaling Specificity.

Highly homologous members of the Gα family, Gα, have distinct tissue distributions and physiological functions, yet the functional properties of these proteins with respect to GDP/GTP binding and regulation of adenylate cyclase are very similar. We recently identified PDZ-RhoGEF (PRG) as a novel Gα effector, however, it is poorly activated by Gα. Here, in a proteomic proximity labeling screen we observed a strong preference for Gα relative to Gα with respect to engagement of a broad range of potential targets.

Phospholipase C-ε defines a PACAP-stimulated pathway for secretion in the chromaffin cell.

Adrenomedullary chromaffin cells respond to splanchnic (sympathetic) nerve stimulation by releasing stress hormones into the circulation. The signal for hormone secretion is encoded in the neurotransmitters - especially acetylcholine (ACh) and pituitary adenylate cyclase activating polypeptide (PACAP) - that are released into the splanchnic-chromaffin cell synapse. However, functional differences in the effects of ACh and PACAP on the chromaffin cell secretory response are not well defined.

A Small Contribution to a Large System: The Leptin Receptor Complex.

Obesity is a classified epidemic, increasing the risk of secondary diseases such as diabetes, inflammation, cardiovascular disease, and cancer. The pleiotropic hormone leptin is the proposed link for the gut-brain axis controlling nutritional status and energy expenditure. Research into leptin signaling provides great promise toward discovering therapeutics for obesity and its related diseases targeting leptin and its cognate leptin receptor (LEP-R).

PACAP and acetylcholine cause distinct Ca2+ signals and secretory responses in chromaffin cells.

The adrenomedullary chromaffin cell transduces chemical messages into outputs that regulate end organ function throughout the periphery. At least two important neurotransmitters are released by innervating preganglionic neurons to stimulate exocytosis in the chromaffin cell-acetylcholine (ACh) and pituitary adenylate cyclase activating polypeptide (PACAP). Although PACAP is widely acknowledged as an important secretagogue in this system, the pathway coupling PACAP stimulation to chromaffin cell secretion is poorly understood.

Phospholipase Cε insufficiency causes ascending aortic aneurysm and dissection.

Phospholipase Cε (PLCε) is a phospholipase C isoform with a wide range of physiological functions. It has been implicated in aortic valve disorders, but its role in frequently associated aortic disease remains unclear. To determine the role of PLCε in thoracic aortic aneurysm and dissection (TAAD) we used PLCε-deficient mice, which develop aortic valve insufficiency and exhibit aortic dilation of the ascending thoracic aorta and arch without histopathological evidence of injury.

Coincident Regulation of PLC Signaling by Gq-Coupled and -Opioid Receptors Opposes Opioid-Mediated Antinociception.

Pain management is an important problem worldwide. The current frontline approach for pain management is the use of opioid analgesics. The primary analgesic target of opioids is the -opioid receptor (MOR).

Subcellular β-Adrenergic Receptor Signaling in Cardiac Physiology and Disease.

Adrenergic receptors are critical regulators of cardiac function with profound effects on cardiac output during sympathetic stimulation. Chronic stimulation of the adrenergic system of the heart under conditions of cardiac stress leads to cardiac dysfunction, hypertrophy, and ultimately failure. Emerging data have revealed that G protein-coupled receptors in intracellular compartments are functionally active and regulate distinct cellular processes from those at the cell surface.

A naturally occurring membrane-anchored Gα variant, XLα, activates phospholipase Cβ4.

Extra-large stimulatory Gα (XLα) is a large variant of G protein α subunit (Gα) that uses an alternative promoter and thus differs from Gα at the first exon. XLα activation by G protein-coupled receptors mediates cAMP generation, similarly to Gα; however, Gα and XLα have been shown to have distinct cellular and physiological functions. For example, previous work suggests that XLα can stimulate inositol phosphate production in renal proximal tubules and thereby regulate serum phosphate levels.

A network of Gα signaling partners is revealed by proximity labeling proteomics analysis and includes PDZ-RhoGEF.

G protein–coupled receptors (GPCRs) that couple to the Gα family of G proteins are key regulators of cell and tissue physiology. Our previous work has revealed new roles for Gα in regulating the migration of neutrophils and fibrosarcoma cells downstream of activated chemoattractant receptors. Here, we used an intact cell proximity–based labeling coupled to tandem mass tag (TMT)–based quantitative proteomics analysis to identify proteins that selectively interacted with the GTP-bound form of Gα.

Uveal melanoma-associated mutations in PLCβ4 are constitutively activating and promote melanocyte proliferation and tumorigenesis.

Activating mutations in Gα proteins are frequent in uveal melanoma, the most common eye cancer arising from the uveal tract. A small proportion of uveal melanomas have a D630Y mutation in phospholipase C β4 (PLCβ4), an effector of Gα. Here, we found that the D630Y mutation in PLCβ4 results in a high level of constitutive PLCβ4 activity.

A universal allosteric mechanism for G protein activation.

G proteins play a central role in signal transduction and pharmacology. Signaling is initiated by cell-surface receptors, which promote guanosine triphosphate (GTP) binding and dissociation of Gα from the Gβγ subunits. Structural studies have revealed the molecular basis of subunit association with receptors, RGS proteins, and downstream effectors.

β-arrestin mediates communication between plasma membrane and intracellular GPCRs to regulate signaling.

It has become increasingly apparent that G protein-coupled receptor (GPCR) localization is a master regulator of cell signaling. However, the molecular mechanisms involved in this process are not well understood. To date, observations of intracellular GPCR activation can be organized into two categories: a dependence on OCT3 cationic channel-permeable ligands or the necessity of endocytic trafficking.

Activation of Phospholipase C β by Gβγ and Gα Involves C-Terminal Rearrangement to Release Autoinhibition.

Phospholipase C (PLC) enzymes hydrolyze phosphoinositide lipids to inositol phosphates and diacylglycerol. Direct activation of PLCβ by Gα and/or Gβγ subunits mediates signaling by Gq and some Gi coupled G-protein-coupled receptors (GPCRs), respectively. PLCβ isoforms contain a unique C-terminal extension, consisting of proximal and distal C-terminal domains (CTDs) separated by a flexible linker.

Hypertension induces glomerulosclerosis in phospholipase C-ε1 deficiency.

Loss-of-function mutations in phospholipase C-ε1 (PLCE1) have been detected in patients with nephrotic syndrome, but other family members with the same mutation were asymptomatic, suggesting additional stressor are required to cause the full phenotype. Consistent with these observations, we determined that global -deficient mice have histologically normal glomeruli and no albuminuria at baseline. Angiotensin II (ANG II) is known to induce glomerular damage in genetically susceptible individuals.