scholarly journals G protein–coupled receptor kinase 2 (GRK2) is localized to centrosomes and mediates epidermal growth factor–promoted centrosomal separation

2013 ◽  
Vol 24 (18) ◽  
pp. 2795-2806 ◽  
Author(s):  
Christopher H. So ◽  
Allison Michal ◽  
Konstantin E. Komolov ◽  
Jiansong Luo ◽  
Jeffrey L. Benovic
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
G Protein ◽  
Growth Factor Receptor ◽  
Dominant Negative ◽  
G Protein Coupled Receptor ◽  
Microtubule Nucleation ◽  
Epidermal Growth ◽  
G Protein Coupled

G protein–coupled receptor kinases (GRKs) play a central role in regulating receptor signaling, but recent studies suggest a broader role in modulating normal cellular functions. For example, GRK5 has been shown to localize to centrosomes and regulate microtubule nucleation and cell cycle progression. Here we demonstrate that GRK2 is also localized to centrosomes, although it has no role in centrosome duplication or microtubule nucleation. Of interest, knockdown of GRK2 inhibits epidermal growth factor receptor (EGFR)–mediated separation of duplicated centrosomes. This EGFR/GRK2-mediated process depends on the protein kinases mammalian STE20-like kinase 2 (Mst2) and Nek2A but does not involve polo-like kinase 1. In vitro analysis and dominant-negative approaches reveal that GRK2 directly phosphorylates and activates Mst2. Collectively these findings demonstrate that GRK2 is localized to centrosomes and plays a central role in mitogen-promoted centrosome separation most likely via its ability to phosphorylate Mst2.

scholarly journals The Ubiquitin-like Protein PLIC-2 Is a Negative Regulator of G Protein-coupled Receptor Endocytosis

2008 ◽  
Vol 19 (3) ◽  
pp. 1252-1260 ◽  
Author(s):  
Elsa-Noah N'Diaye ◽  
Aylin C. Hanyaloglu ◽  
Kimberly K. Kajihara ◽  
Manojkumar A. Puthenveedu ◽  
Ping Wu ◽  
...  
Keyword(s):  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
G Protein ◽  
Growth Factor Receptor ◽  
Negative Regulator ◽  
Regulatory Function ◽  
Receptor Endocytosis ◽  
Epidermal Growth ◽  
G Protein Coupled

The activity of many signaling receptors is regulated by their endocytosis via clathrin-coated pits (CCPs). For G protein-coupled receptors (GPCRs), recruitment of the adaptor protein arrestin to activated receptors is thought to be sufficient to drive GPCR clustering in CCPs and subsequent endocytosis. We have identified an unprecedented role for the ubiquitin-like protein PLIC-2 as a negative regulator of GPCR endocytosis. Protein Linking IAP to Cytoskeleton (PLIC)-2 overexpression delayed ligand-induced endocytosis of two GPCRs: the V2 vasopressin receptor and β-2 adrenergic receptor, without affecting endocytosis of the transferrin or epidermal growth factor receptor. The closely related isoform PLIC-1 did not affect receptor endocytosis. PLIC-2 specifically inhibited GPCR concentration in CCPs, without affecting membrane recruitment of arrestin-3 to activated receptors or its cellular levels. Depletion of cellular PLIC-2 accelerated GPCR endocytosis, confirming its regulatory function at endogenous levels. The ubiquitin-like domain of PLIC-2, a ligand for ubiquitin-interacting motifs (UIMs), was required for endocytic inhibition. Interestingly, the UIM-containing endocytic adaptors epidermal growth factor receptor protein substrate 15 and Epsin exhibited preferential binding to PLIC-2 over PLIC-1. This differential interaction may underlie PLIC-2 specific effect on GPCR endocytosis. Identification of a negative regulator of GPCR clustering reveals a new function of ubiquitin-like proteins and highlights a cellular requirement for exquisite regulation of receptor dynamics.

scholarly journals Estrogen Action Via the G Protein-Coupled Receptor, GPR30: Stimulation of Adenylyl Cyclase and cAMP-Mediated Attenuation of the Epidermal Growth Factor Receptor-to-MAPK Signaling Axis

2002 ◽  
Vol 16 (1) ◽  
pp. 70-84 ◽  
Author(s):  
Edward J. Filardo ◽  
Jeffrey A. Quinn ◽  
A. Raymond Frackelton ◽  
Kirby I. Bland
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Adenylyl Cyclase ◽  
G Protein ◽  
Egf Receptor ◽  
G Protein Coupled Receptor ◽  
Estrogen Action ◽  
Epidermal Growth ◽  
G Protein Coupled ◽  
Stimulation Of

Abstract Estrogen triggers rapid yet transient activation of the MAPKs, extracellular signal-regulated kinase (Erk)-1 and Erk-2. We have reported that this estrogen action requires the G protein-coupled receptor, GPR30, and occurs via Gβγ-subunit protein-dependent transactivation of the epidermal growth factor (EGF) receptor through the release of pro-heparan-bound EGF from the cell surface. Here we investigate the mechanism by which Erk-1/-2 activity is rapidly restored to basal levels after estrogen stimulation. Evidence is provided that attenuation of Erk-1/-2 activity by estrogen occurs via GPR30-dependent stimulation of adenylyl cyclase and cAMP-dependent signaling that results in Raf-1 inactivation. We show that 17β-E2 represses EGF-induced activation of the Raf-to-Erk pathway in human breast carcinoma cells that express GPR30, including MCF-7 and SKBR3 cells which express both or neither, ER, respectively. MDA-MB-231 cells, which express ERβ, but not ERα, and low levels of GPR30 protein, are unable to stimulate adenylyl cyclase or promote estrogen-mediated blockade of EGF-induced activation of Erk-1/-2. Pretreatment of MDA-MB-231 cells with cholera toxin, which ADP-ribosylates and activates Gαs subunit proteins, results in G protein-coupled receptor (GPCR)-independent adenylyl cyclase activity and suppression of EGF-induced Erk-1/-2 activity. Transfection of GPR30 into MDA-MB-231 cells restores their ability to stimulate adenylyl cyclase and attenuate EGF-induced activation of Erk-1/-2 by estrogen. Moreover, GPR30-dependent, cAMP-mediated attenuation of EGF-induced Erk-1/-2 activity was achieved by ER antagonists such as tamoxifen or ICI 182, 780; yet not by 17α-E2 or progesterone. Thus, our data delineate a novel mechanism, requiring GPR30 and estrogen, that acts to regulate Erk-1/-2 activity via an inhibitory signal mediated by cAMP. Coupled with our prior findings, these current data imply that estrogen balances Erk-1/-2 activity through a single GPCR via two distinct G protein-dependent signaling pathways that have opposing effects on the EGF receptor-to-MAPK pathway.

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