G protein-regulated endocytic trafficking of adenylyl cyclase type 9.

André M Lazar,Asuka Inoue,Aparna B Sundaram,Mark Von Zastrow,Carmen W Dessauer,J Silvio Gutkind,Tanya A Baldwin,Roshanak Irannejad

doi:10.7554/elife.58039

Abstract

GPCRs are increasingly recognized to initiate signaling via heterotrimeric G proteins as they move through the endocytic network, but little is known about how relevant G protein effectors are localized. Here we report selective trafficking of adenylyl cyclase type 9 (AC9) from the plasma membrane to endosomes while adenylyl cyclase type 1 (AC1) remains in the plasma membrane, and stimulation of AC9 trafficking by ligand-induced activation of Gs-coupled GPCRs. AC9 transits a similar, dynamin-dependent early endocytic pathway as ligand-activated GPCRs. However, unlike GPCR traffic control which requires β-arrestin but not Gs, AC9 traffic control requires Gs but not β-arrestin. We also show that AC9, but not AC1, mediates cAMP production stimulated by endogenous receptor activation in endosomes. These results reveal dynamic and isoform-specific trafficking of adenylyl cyclase in the endocytic network, and a discrete role of a heterotrimeric G protein in regulating the subcellular distribution of a relevant effector.

Highlights

G protein-coupled receptors (GPCRs) comprise nature’s largest family of signaling receptors and an important class of therapeutic targets (Lefkowitz, 2007; Rosenbaum et al, 2009)
adenylyl cyclase type 9 (AC9) labeled in its C-terminal cytoplasmic domain with GFP (AC9-GFP) redistributed, enabling live-cell confocal imaging which revealed mobile AC9-containing endosomes (Video 1). These results indicate that AC9 traffics dynamically to endosomes containing beta-2 adrenergic receptor (b2AR), this trafficking is isoform-specific because adenylyl cyclase type 1 (AC1) remains in the plasma membrane, and it is regulated because AC9 accumulation in endosomes is increased by b2AR activation
This compartment is known to accumulate a wide variety of GPCRs, and it has been explicitly shown to be a site of Gs activation by the b2AR (Irannejad et al, 2013)

Summary

Introduction

G protein-coupled receptors (GPCRs) comprise nature’s largest family of signaling receptors and an important class of therapeutic targets (Lefkowitz, 2007; Rosenbaum et al, 2009). GPCRs are sonamed because a major mechanism by which they mediate transmembrane signaling is through ligand-dependent activation of heterotrimeric G proteins that act as intracellular signal transducers (Gilman, 1987; Hilger et al, 2018; Spiegel, 1987; Stryer and Bourne, 1986; Sunahara et al, 1996) This conserved signaling cascade invariably requires one additional component, an ‘effector’ protein which is regulated by the G protein to convey the signal downstream (Dessauer et al, 1996; Gilman, 1987; Rosenbaum et al, 2009).

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