Expression of a β-adrenergic receptor kinase 1 inhibitor prevents the development of myocardial failure in gene-targeted mice

Abstract

Heart failure is accompanied by severely impaired ␤-adrenergic receptor (␤AR) function, which includes loss of ␤AR density and functional uncoupling of remaining receptors. An important mechanism for the rapid desensitization of ␤AR function is agonist-stimulated receptor phosphorylation by the ␤AR kinase (␤ARK1), an enzyme known to be elevated in failing human heart tissue. To investigate whether alterations in ␤AR function contribute to the development of myocardial failure, transgenic mice with cardiac-restricted overexpression of either a peptide inhibitor of ␤ARK1 or the ␤ 2 AR were mated into a genetic model of murine heart failure (MLP ؊͞؊ ). In vivo cardiac function was assessed by echocardiography and cardiac catheterization. Both MLP ؊͞؊ and MLP ؊͞؊ ͞␤ 2 AR mice had enlarged left ventricular (LV) chambers with significantly reduced fractional shortening and mean velocity of circumferential fiber shortening. In contrast, MLP ؊͞؊ ͞␤ARKct mice had normal LV chamber size and function. Basal LV contractility in the MLP ؊͞؊ ͞␤ARKct mice, as measured by LV dP͞dtmax, was increased significantly compared with the MLP ؊͞؊ mice but less than controls. Importantly, heightened ␤AR desensitization in the MLP ؊͞؊ mice, measured in vivo (responsiveness to isoproterenol) and in vitro (isoproterenol-stimulated membrane adenylyl cyclase activity), was completely reversed with overexpression of the ␤ARK1 inhibitor. We report here the striking finding that overexpression of this inhibitor prevents the development of cardiomyopathy in this murine model of heart failure. These findings implicate abnormal ␤AR-G protein coupling in the pathogenesis of the failing heart and point the way toward development of agents to inhibit ␤ARK1 as a novel mode of therapy.One of the most important mechanisms for rapidly regulating ␤-adrenergic receptor (␤AR) function is agonist-stimulated receptor phosphorylation by G protein-coupled receptor kinases (GRKs) resulting in decreased sensitivity to further catecholamine stimulation (1, 2). ␤ARK1 is a member of the multigene GRK family that regulates a wide variety of receptors that couple to heterotrimeric G proteins (1, 2). Desensitization of agonistoccupied receptors by the cytosolic ␤AR kinase (␤ARK1) requires a membrane-targeting event before its activation and receptor phosphorylation, which is mediated by a direct physical interaction between residues within the carboxyl terminus of ␤ARK1 and the dissociated membrane-anchored ␤␥ subunits of G proteins (G␤␥) (3, 4).Heart failure is a disease characterized by left ventricular (LV) dysfunction associated with a complex of symptoms that relate to inadequate perfusion of tissues and pulmonary congestion. Although the fundamental molecular abnormality that causes this progressive deterioration in cardiac function is unknown, one of the leading candidates is abnormal ␤AR signaling. Chronic human heart failure is characterized by severely attenuated ␤AR signaling, resulting from diminished receptor number and impaired receptor function (5...

Effects of two Gβγ-binding proteins – N-terminally truncated phosducin and β-adrenergic receptor kinase C terminus (βARKct) – in heart failure

“…5 Also the finding of improved contractility after gene transfer of Ad-bARKct-GFP in vivo corresponds well with the improvement of LV function and outcome measured in similar disease models 5 or in cardiacspecific bARKct-transgenic mice. 5,9,8 To a similar extent as bARKct, also overexpression of nt-del-phosducin enhanced basal contraction and maximal contractility of both normal or failing cardiomyocytes. Moreover, a clear leftward shift of the concentration-contractility curve occurred ( Figure 6).…”

“…Overexpression of bARKct in several models of heart failure has been shown to result in improved hemodynamic conditions, for example, after somatic gene transfer into rabbits with heart failure because of rapid pacing 5 or myocardial infarction. 6,7 Also, crossbreeding of cardiac-specific bARKct-transgenic mice with mouse models of heart failure due to ablation of the muscle LIM protein (MLP (À/À) mice), 8 due to a myosin heavy chain mutant 9 or due to overexpression of calsequestrin 10 prevented the development of heart failure or improved survival. This effect was additive to the treatment of heart failure with b-blockers.…”

Rescue of Contractile Parameters and Myocyte Hypertrophy in Calsequestrin Overexpressing Myocardium by Phospholamban Ablation

“…Cardiac-specific overexpression of the ␤-adrenergic receptor kinase inhibitor was also reported to improve cardiac function in the same model of cardiomyopathy (51). Since phospholamban is the major substrate in the cardiac ␤-adrenergic pathway, it is interesting to propose that at least a part of the beneficial effects by the latter genetic manipulation may be mediated by the phosphorylation level of phospholamban.…”

Regulation of cardiac contractility by Rab4-modulated β2-adrenergic receptor recycling

“…␤AR move from plasma membranes into endosomal structures where they are either targeted for lysosomal degradation or recycled as fully functional receptors by vesicular trafficking back to plasma membranes (resensitization) (10). Although the roles for GRK-mediated ␤AR receptoreffector uncoupling in cardiac health and disease have been established (11)(12)(13)(14), the pathophysiological roles of modulated ␤AR receptor internalization͞recycling have not been defined, largely because suitable in vivo experimental models do not exist. Herein, we used Rab-GTPases, regulators of inter-vesicular membrane transport (15), to define effects of modulated ␤AR trafficking in the in vivo heart.…”