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We have recently demonstrated that the amino-terminal domain of G protein coupled receptor kinase (GRK) type 5, (GRK5-NT) inhibits NFκB activity in cardiac cells leading to a significant amelioration of LVH. Since GRK5-NT is known to bind calmodulin, this study aimed to evaluate the functional role of GRK5-NT in the regulation of calcium-calmodulin-dependent transcription factors. We found that the overexpression of GRK5-NT in cardiomyoblasts significantly reduced the activation and the nuclear translocation of NFAT and its cofactor GATA-4 in response to phenylephrine (PE). These results were confirmed in vivo in spontaneously hypertensive rats (SHR), in which intramyocardial adenovirus-mediated gene transfer of GRK5-NT reduced both wall thickness and ventricular mass by modulating NFAT and GATA-4 activity. To further verify in vitro the contribution of calmodulin in linking GRK5-NT to the NFAT/GATA-4 pathway, we examined the effects of a mutant of GRK5 (GRK5-NTPB), which is not able to bind calmodulin. When compared to GRK5-NT, GRK5-NTPB did not modify PE-induced NFAT and GATA-4 activation. In conclusion, this study identifies a double effect of GRK5-NT in the inhibition of LVH that is based on the regulation of multiple transcription factors through means of different mechanisms and proposes the amino-terminal sequence of GRK5 as a useful prototype for therapeutic purposes. [ABSTRACT FROM AUTHOR]

G protein-coupled receptors (GPCRs) are cell surface receptors that respond to a wide variety of stimuli, from light, odorants, hormones, and neurotransmitters to proteins and extracellular calcium. GPCRs represent the largest family of signaling proteins targeted by many clinically used drugs. Recent studies shed light on the conformational changes that accompany GPCR activation and the structural state of the receptor necessary for the interactions with the three classes of proteins that preferentially bind active GPCRs, G proteins, G protein-coupled receptor kinases (GRKs), and arrestins. Importantly, structural and biophysical studies also revealed activation-related conformational changes in these three types of signal transducers. Here, we summarize what is already known and point out questions that still need to be answered. Clear understanding of the structural basis of signaling by GPCRs and their interaction partners would pave the way to designing signaling-biased proteins with scientific and therapeutic potential. [ABSTRACT FROM AUTHOR]

Cross-talk between the sympathetic nervous system (SNS) and immune system is vital for health and well-being. Infection, tissue injury and inflammation raise firing rates of sympathetic nerves, increasing their release of norepinephrine (NE) in lymphoid organs and tissues. NE stimulation of β2-adrenergic receptors (ARs) in immune cells activates the cAMP-protein kinase A (PKA) intracellular signaling pathway, a pathway that interfaces with other signaling pathways that regulate proliferation, differentiation, maturation and effector functions in immune cells. Immune-SNS cross-talk is required to maintain homeostasis under normal conditions, to develop an immune response of appropriate magnitude after injury or immune challenge, and subsequently restore homeostasis. Typically, β2-AR-induced cAMP is immunosuppressive. However, many studies report actions of β2-AR stimulation in immune cells that are inconsistent with typical cAMP-PKA signal transduction. Research during the last decade in non-immune organs, has unveiled novel alternative signaling mechanisms induced by β2-AR activation, such as a signaling switch from cAMP-PKA to mitogen-activated protein kinase (MAPK) pathways. If alternative signaling occurs in immune cells, it may explain inconsistent findings of sympathetic regulation of immune function. Here, we review β2-AR signaling, assess the available evidence for alternative signaling in immune cells, and provide insight into the circumstances necessary for "signal switching" in immune cells. [ABSTRACT FROM AUTHOR]

The lysophosphatidic acid 3 receptor (LPA3) participates in different physiological actions and in the pathogenesis of many diseases through the activation of different signal pathways. Knowledge of the regulation of the function of the LPA3 receptor is a crucial element for defining its roles in health and disease. This review describes what is known about the signaling pathways activated in terms of its various actions. Next, we review knowledge on the structure of the LPA3 receptor, the domains found, and the roles that the latter might play in ligand recognition, signaling, and cellular localization. Currently, there is some information on the action of LPA3 in different cells and whole organisms, but very little is known about the regulation of its function. Areas in which there is a gap in our knowledge are indicated in order to further stimulate experimental work on this receptor and on other members of the LPA receptor family. We are convinced that knowledge on how this receptor is activated, the signaling pathways employed and how the receptor internalization and desensitization are controlled will help design new therapeutic interventions for treating diseases in which the LPA3 receptor is implicated. [ABSTRACT FROM AUTHOR]

Investigations on functional selectivity of GPCR ligands have become increasingly important to identify compounds with a potentially more beneficial side effect profile. In order to discriminate between individual signaling pathways, the determination of β-arrestin2 recruitment, in addition to G-protein activation, is of great value. In this study, we established a sensitive split luciferase-based assay with the ability to quantify β-arrestin2 recruitment to D 2long and D 3 receptors and measure time-resolved β-arrestin2 recruitment to the D 2long receptor after agonist stimulation. We were able to characterize several standard (inverse) agonists as well as antagonists at the D 2long R and D 3 R subtypes, whereas for the D 4.4 R, no β-arrestin2 recruitment was detected, confirming previous reports. Extensive radioligand binding studies and comparisons with the respective wild-type receptors confirm that the attachment of the Emerald luciferase fragment to the receptors does not affect the integrity of the receptor proteins. Studies on the involvement of GRK2/3 and PKC on the β-arrestin recruitment to the D 2long R and D 3 R, as well as at the D 1 R using different kinase inhibitors, showed that the assay could also contribute to the elucidation of signaling mechanisms. Its broad applicability, which provides concentration-dependent and kinetic information on receptor/β-arrestin2 interactions, renders this homogeneous assay a valuable method for the identification of biased agonists.

Inflammation is a key mechanism of cardiovascular diseases. It is an essential component of atherosclerosis and a significant risk factor for the development of cardiovascular events. In the crosstalk between inflammation and cardiovascular diseases, the transcription factor NFκB seems to be a key player since it is involved in the development and progression of both inflammation and cardiac and vascular damage. In this review, we deal with the recent findings of the role of inflammation in cardiac diseases, focusing, in particular, on NFκB as a functional link. We describe strategies for the therapeutic targeting of NFκB as a potential strategy for the failing heart. [ABSTRACT FROM AUTHOR]