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

Abstract

Catecholaminergic activation of myocardial ␤-adrenergic receptors (␤AR) is the principle mechanism regulating cardiac function. Agonists desensitize ␤AR through G protein-coupled receptor kinase-mediated uncoupling and ␤-arrestin-mediated internalization. Although inhibition of myocardial G protein-coupled receptor kinase-2 enhances cardiac function and reverses heart failure, pathophysiological effects of modulated ␤AR internalization͞re-cycling are unknown. We used mutation and transgenic expression of Rab4, which regulates vesicular transport of heptahelical receptors to plasma membranes, to interrogate in vivo ␤AR trafficking and cardiac function. Expression of constitutively active Rab4 Q72L had no effects on cardiac structure or function, but dominant inhibitor Rab4 S27N impaired responsiveness to endogenous and exogenous catecholamines. To relate ␤AR trafficking to diminished cardiac function, Rab4 mutant mice were crossbred with mice overexpressing human ␤2AR. In unstimulated ␤2AR overexpressors, ␤2AR localized to heavier endosomes and translocated to lighter, caveolin-rich fractions after isoproterenol stimulation. Coexpression of ␤2AR with activated Rab4 Q72L caused loss of receptors from heavier endosomes while retaining normal inotropy. In contrast, coexpression of ␤2AR with inhibitory Rab4 S27N mimicked isoproterenol-induced receptor redistribution to caveolae, with diminished cardiac inotropy. Rab4 inhibition alone prevented resensitization after isoproterenol-induced in vivo adrenergic desensitization. Confocal and ultrastructural analyses revealed bizarre vesicular structures and abnormal accumulation of ␤2AR in the sarcoplasm and subsarcollema of Rab4 S27N, but not Q72L, mice. These data provide evidence for constant bidirectional sarcollemal-vesicular ␤AR trafficking in the in vivo heart and show that Rab4-mediated recycling of internalized ␤AR is necessary for normal cardiac catecholamine responsiveness and resensitization after agonist exposure.

Enhanced Hypotensive, Bradycardic, and Hypnotic Responses to α2-Adrenergic Agonists in Spinophilin-Null Mice Are Accompanied by Increased G Protein Coupling to the α2A-Adrenergic Receptor

“…Such a role for receptor endocytosis/recycling for signaling has been observed in other systems, as well. For example, clathrin-mediated internalization and receptor recycling has been shown to be essential for sustained EGFR signaling (Sigismund et al, 2008), and recycling of ␤AR is required for sustained cardiac responsiveness to catecholamine in vivo (Odley et al, 2004). Furthermore, the endosome can serve as a intracellular platform for assembly of signaling complex, either through receptor-associated scaffolding proteins such as arrestin (Violin and Lefkowitz, 2007) or by recruiting signaling molecules to endosomal resident proteins such as Rab5 (Zoncu et al, 2009).…”

Restoration of β-Adrenergic Receptor Signaling and Contractile Function in Heart Failure by Disruption of the βARK1/Phosphoinositide 3-Kinase Complex

“…25 Our recent in vitro studies have shown that generation of phosphoinositides by PI3K at the site of activated receptors is required for efficient internalization of ␤ARs after agonist stimulation. 9 Consistently, in pigs with pacing-induced heart failure, ␤AR redistribution to intracellular compartments was accompanied by a significant increase in membrane-targeted PI3K activity.…”

Dynamic of Ion Channel Expression at the Plasma Membrane of Cardiomyocytes

“…Rab9 is involved in transport from late endosome to the TGN (243), and Rab7 is also associated with the late endosome but instead directs the trafficking of its contents to the lysosome (120,337) and to the proteasome (102) for degradation. All of these Rab GTPases are expressed in cardiomyocytes (125,197,239,311).…”