Enhancement of the Recycling and Activation of β-Adrenergic Receptor by Rab4 GTPase in Cardiac Myocytes

Catalin M Filipeanu,Fuguo Zhou,May L Lam,Kenneth E Kerut,William C Claycomb,Guangyu Wu

doi:10.1074/jbc.m511460200

Catalin M Filipeanu, Fuguo Zhou + Show 4 more

Open Access

https://doi.org/10.1074/jbc.m511460200

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Abstract

We investigate the role of Rab4, a Ras-like small GTPase coordinating protein transport from the endosome to the plasma membrane, on the recycling and activation of endogenous beta-adrenergic receptor (beta-AR) in HL-1 cardiac myocytes in vitro and transgenic mouse hearts in vivo. Beta1-AR, the predominant subtype of beta-AR in HL-1 cardiac myocytes, was internalized after stimulation with isoproterenol (ISO) and fully recycled at 4 h upon ISO removal. Transient expression of Rab4 markedly facilitated recycling of internalized beta-AR to the cell surface and enhanced beta-AR signaling as measured by ISO-stimulated cAMP production. Transgenic overexpression of Rab4 in the mouse myocardium significantly increased the number of beta-AR in the plasma membrane and augmented cAMP production at the basal level and in response to ISO stimulation. Rab4 overexpression induced concentric cardiac hypertrophy with a moderate increase in ventricle/body weight ratio and posterior wall thickness and a selective up-regulation of the beta-myosin heavy chain gene. These data provide the first evidence indicating that Rab4 is a rate-limiting factor for the recycling of endogenous beta-AR and augmentation of Rab4-mediated traffic enhances beta-AR function in cardiac myocytes.

Highlights

The number of ␤-Adrenergic receptors (ARs) at the cell surface at a certain time determines the amplitude of functional response to ␤-AR stimulation by extracellular hormones
Several studies have demonstrated that co-expression of the dominant-negative GDP-bound Rab4 mutant inhibits G protein-coupled receptor (GPCR) transport from the endosome to the plasma membrane after stimulation with their agonists, suggesting that normal Rab4 function is required for the recycling of internalized receptors (9, 18 –20)
We investigated the influence of increased Rab4 function by overexpressing wildtype Rab4 on the transport of endogenous ␤-AR to the plasma membrane in both cardiac mycoytes in vitro and mouse hearts in vivo

Summary

Introduction

The number of ␤-AR at the cell surface at a certain time determines the amplitude of functional response to ␤-AR stimulation by extracellular hormones. Our data demonstrated that increased wild-type Rab4 expression facilitated recycling to the plasma membrane and signaling of ␤-AR in cultured HL-1 cardiac myocytes. To determine whether increased cAMP production in Rab4-transfected HL-1 myocytes is due to the alteration of adenylyl cyclase activity, we measured cAMP production in response to stimulation with forskolin, which directly activates adenylyl cyclases, bypassing the plasma membrane receptors.

Results

Conclusion