Faculty Opinions recommendation of Myocardin-Related Transcription Factor A and Yes-Associated Protein Exert Dual Control in G Protein-Coupled Receptor- and RhoA-Mediated Transcriptional Regulation and Cell Proliferation.

Leucine-rich repeat-containing G-protein-coupled receptor 8 (LGR8, or RXFP2) is a member of the type C leucine-rich repeat-containing G protein-coupled receptor family, and its endogenous ligand is insulin-like peptide-3 (INSL3). Although LGR8 expression has been demonstrated in various human tissues, including testis, ovary, brain and kidney, the precise roles of this receptor in many of these tissues are unknown. In an effort to better understand INSL3–LGR8 systems in the rat, we cloned the full-length Lgr8 cDNA and investigated the presence and cellular localization of Lgr8 mRNA expression in adult and developing rat kidney. On the basis of these findings, we investigated the presence and distribution of renal 125I-labelled human INSL3-binding sites and the nature of INSL3–LGR8 signalling in cultured renal cells. Thus, using in situ hybridization histochemistry, cells expressing Lgr8 mRNA were observed in glomeruli of renal cortex from adult rats and were tentatively identified as mesangial cells. Quantitative, real-time PCR analysis of the developmental profile of Lgr8 mRNA expression in kidney revealed highest relative levels at late stage gestation (embryonic day 18), with a sharp decrease after birth and lowest levels in the adult. During development, silver grains associated with Lgr8 mRNA hybridization were observed overlying putative mesangial cells in mature glomeruli, with little or no signal associated with less-mature glomeruli. In adult and developing kidney, specific 125I-INSL3-binding sites were associated with glomeruli throughout the renal cortex. In primary cultures of glomerular cells, synthetic human INSL3 specifically and dose-dependently inhibited cell proliferation over a 48 h period, further suggesting the presence of functional LGR8 (receptors) on these cells (mesangial and others). These findings suggest INSL3–LGR8 signalling may be involved in the genesis and/or developmental maturation of renal glomeruli and possibly in regulating mesangial cell density in adult rat kidney.

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Activation of Intracellular Signaling Pathways by the Murine Cytomegalovirus G Protein-Coupled Receptor M33 Occurs via PLC-β/PKC-Dependent and -Independent Mechanisms

Journal of Virology ◽

10.1128/jvi.02116-08 ◽

2009 ◽

Vol 83 (16) ◽

pp. 8141-8152 ◽

Cited By ~ 15

Author(s):

Joseph D. Sherrill ◽

Melissa P. Stropes ◽

Olivia D. Schneider ◽

Diana E. Koch ◽

Fabiola M. Bittencourt ◽

...

Keyword(s):

Transcription Factor ◽

Protein Kinase ◽

Signaling Pathways ◽

G Protein ◽

Intracellular Signaling ◽

Murine Cytomegalovirus ◽

Dependent Manner ◽

G Protein Coupled Receptor ◽

G Protein Coupled

ABSTRACT The presence of numerous G protein-coupled receptor (GPCR) homologs within the herpesvirus genomes suggests an essential role for these genes in viral replication in the infected host. Such is the case for murine cytomegalovirus (MCMV), where deletion of the M33 GPCR or replacement of M33 with a signaling defective mutant has been shown to severely attenuate replication in vivo. In the present study we utilized a genetically altered version of M33 (termed R131A) in combination with pharmacological inhibitors to further characterize the mechanisms by which M33 activates downstream signaling pathways. This R131A mutant of M33 fails to support salivary gland replication in vivo and, as such, is an important tool that can be used to examine the signaling activities of M33. We show that M33 stimulates the transcription factor CREB via heterotrimeric Gq/11 proteins and not through promiscuous coupling of M33 to the Gs pathway. Using inhibitors of signaling molecules downstream of Gq/11, we demonstrate that M33 stimulates CREB transcriptional activity in a phospholipase C-β and protein kinase C (PKC)-dependent manner. Finally, utilizing wild-type and R131A versions of M33, we show that M33-mediated activation of other signaling nodes, including the mitogen-activated protein kinase family member p38α and transcription factor NF-κB, occurs in the absence of Gq/11 and PKC signaling. The results from the present study indicate that M33 utilizes multiple mechanisms to modulate intracellular signaling cascades and suggest that signaling through PLC-β and PKC plays a central role in MCMV pathogenesis in vivo.

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Faculty Opinions recommendation of Distinct ADAM metalloproteinases regulate G protein-coupled receptor-induced cell proliferation and survival.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1021707.246679 ◽

2004 ◽

Author(s):

Judith White

Keyword(s):

Cell Proliferation ◽

G Protein ◽

G Protein Coupled Receptor ◽

G Protein Coupled

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The G Protein-Coupled Receptor GPR30 Inhibits Human Urothelial Cell Proliferation

Endocrinology ◽

10.1210/en.2007-1669 ◽

2008 ◽

Vol 149 (8) ◽

pp. 4024-4034 ◽

Cited By ~ 48

Author(s):

Jian Teng ◽

Zun-Yi Wang ◽

Eric R. Prossnitz ◽

Dale E. Bjorling

Keyword(s):

Cell Proliferation ◽

Cyclin D1 ◽

G Protein ◽

Small Interfering Rna ◽

Urothelial Cell ◽

Urothelial Cells ◽

G Protein Coupled Receptor ◽

17Β Estradiol ◽

Interfering Rna ◽

G Protein Coupled

We have previously shown that estrogen stimulates cell proliferation in both normal and transformed urothelial cells mainly through activation of the two primary estrogen receptors (ERs), ERα and ERβ. A growing body of evidence suggests that estrogen also initiates nongenomic effects that cannot be explained by activation of primary ERs. In the present study, we observed that urothelial cells express high amounts of GPR30, a G protein-coupled receptor recently identified as a candidate for membrane-associated estrogen binding. Membrane- impermeable bovine serum albumin-conjugated 17β-estradiol and the specific GPR30 agonist G-1 both inhibited urothelial cell proliferation in a concentration-dependent manner. Transient overexpression of GPR30 inhibited 17β-estradiol (E2)-induced cell proliferation. Decreased GPR30 expression caused by specific small interfering RNA increased E2-induced cell proliferation. These results indicate that membrane-associated inhibitory effects of E2 on cell proliferation correlate with abundance of GPR30. Although E2 induced a significant increase in caspase-3/7 activity, G-1 did not, suggesting that the GPR30-mediated inhibitory effect on cell proliferation was not caused by apoptosis. Furthermore, we found that G-1 failed to induce c-fos, c-jun, and cyclin D1 expression, and GPR30 overexpression abolished E2-induced c-fos, c-jun, and cyclin D1 expression. However, inactivation of GPR30 by small interfering RNA increased c-fos, c-jun, and cyclin D1 expression. These results suggest that GPR30-mediated inhibition of urothelial cell proliferation is the result of decreased cyclin D1 by down-regulation of activation protein-1 signaling.

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The G Protein–Coupled Receptor GPR30 Mediates the Nontranscriptional Effect of Estrogen on the Activation of PI3K/Akt Pathway in Endometrial Cancer Cells

International Journal of Gynecological Cancer ◽

10.1097/igc.0b013e31827912b8 ◽

2013 ◽

Vol 23 (1) ◽

pp. 52-59 ◽

Cited By ~ 32

Author(s):

Xin Ge ◽

Ruixia Guo ◽

Yuhuan Qiao ◽

Yancai Zhang ◽

Jia Lei ◽

...

Keyword(s):

Cell Proliferation ◽

Endometrial Cancer ◽

Cancer Cells ◽

G Protein ◽

Akt Pathway ◽

G Protein Coupled Receptor ◽

Down Regulation ◽

Phosphorylated Akt ◽

Ishikawa Cells ◽

G Protein Coupled

ObjectiveThe goal of this study was to investigate the effect of G protein–coupled receptor 30 (GPR30) on the activation of PI3K/Akt pathway induced by E2 in endometrial cancer cells.Methods and materialsImmunohistochemistry was performed to determine the location and expression of GPR30, estrogen receptors (ERs), Akt, and phosphorylated Akt. We also investigated the expression of GPR30, ERs, and the level of phosphorylation of Akt induced by E2 in endometrial cancer cells, Ishikawa cells, and HEC-1A cells. We down-regulated the expression of GPR30 in endometrial cancer cell lines by transfection with shGPR30-pGFP-V-RS, a GPR30 antisense expression vector. The cells were then subjected to a proliferation assay. Immunoprecipitation assay was performed to determine whether GPR30 directly bind to PI3K. The stable transfected cells resuspension of 100 μL (5 × 106 cells) was injected subcutaneously into the right flank of athymic mice to perform xenograft tumor formation assays.ResultsE2 stimulated cell proliferation and induced GPR30 expression and PI3K/Akt pathway activation in endometrial cancer cells, Ishikawa cells, and HEC-1A cells, whereas the expression of ERs remained unchangeable. Down-regulation of GPR30 decreased the phosphorylation of Akt and reduced cell proliferation, and GPR30 did not bind to PI3K. Down-regulation of GPR30 significantly inhibited the tumor growth of HEC-1A cells in athymic nude mice.ConclusionsThese findings suggest that GPR30 mediates the nontranscriptional effect of estrogen on the activation of PI3K/Akt pathway in endometrial cancer cells.

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Evidence for Osteocalcin Binding and Activation of GPRC6A in β-Cells

Endocrinology ◽

10.1210/en.2015-2010 ◽

2016 ◽

Vol 157 (5) ◽

pp. 1866-1880 ◽

Cited By ~ 56

Author(s):

Min Pi ◽

Karan Kapoor ◽

Ruisong Ye ◽

Satoru Kenneth Nishimoto ◽

Jeremy C. Smith ◽

...

Keyword(s):

Cell Proliferation ◽

G Protein ◽

Computational Models ◽

Transmembrane Domain ◽

Structural Basis ◽

G Protein Coupled Receptor ◽

Β Cells ◽

Β Cell ◽

Cell Functions ◽

G Protein Coupled

Abstract The possibility that G protein-coupled receptor family C member A (GPRC6A) is the osteocalcin (Ocn)-sensing G protein-coupled receptor that directly regulates pancreatic β-cell functions is controversial. In the current study, we found that Ocn and an Ocn-derived C-terminal hexapeptide directly activate GPRC6A-dependent ERK signaling in vitro. Computational models probe the structural basis of Ocn binding to GPRC6A and predict that the C-terminal hexapeptide docks to the extracellular side of the transmembrane domain of GPRC6A. Consistent with the modeling, mutations in the computationally identified binding pocket of GPRC6A reduced Ocn and C-terminal hexapeptide activation of this receptor. In addition, selective deletion of Gprc6a in β-cells (Gprc6aβ-cell-cko) by crossing Gprc6aflox/flox mice with Ins2-Cre mice resulted in reduced pancreatic weight, islet number, insulin protein content, and insulin message expression. Both islet size and β-cell proliferation were reduced in Gprc6aβ-cell-cko compared with control mice. Gprc6aβ-cell-cko exhibited abnormal glucose tolerance, but normal insulin sensitivity. Islets isolated from Gprc6aβ-cell-cko mice showed reduced insulin simulation index in response to Ocn. These data establish the structural basis for Ocn direct activation of GPRC6A and confirm a role for GPRC6A in regulating β-cell proliferation and insulin secretion.

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