scholarly journals GPR-4 Is a Predicted G-Protein-Coupled Receptor Required for Carbon Source-Dependent Asexual Growth and Development in Neurospora crassa

2006 ◽  
Vol 5 (8) ◽  
pp. 1287-1300 ◽  
Author(s):  
Liande Li ◽  
Katherine A. Borkovich
Keyword(s):  
Carbon Source ◽  
Neurospora Crassa ◽  
G Protein ◽  
Growth And Development ◽  
Carbon Sources ◽  
Wild Type ◽  
G Protein Coupled Receptor ◽  
G Protein Coupled ◽  
Camp Levels ◽  
Mass Accumulation

ABSTRACT The filamentous fungus Neurospora crassa is able to utilize a wide variety of carbon sources. Here, we examine the involvement of a predicted G-protein-coupled receptor (GPCR), GPR-4, during growth and development in the presence of different carbon sources in N. crassa. Δgpr-4 mutants have reduced mass accumulation compared to the wild type when cultured on high levels of glycerol, mannitol, or arabinose. The defect is most severe on glycerol and is cell density dependent. The genetic and physical relationship between GPR-4 and the three N. crassa Gα subunits (GNA-1, GNA-2, and GNA-3) was explored. All three Gα mutants are defective in mass accumulation when cultured on glycerol. However, the phenotypes of Δgna-1 and Δgpr-4 Δgna-1 mutants are identical, introduction of a constitutively activated gna-1 allele suppresses the defects of the Δgpr-4 mutation, and the carboxy terminus of GPR-4 interacts most strongly with GNA-1 in the yeast two-hybrid assay. Although steady-state cyclic AMP (cAMP) levels are normal in Δgpr-4 strains, exogenous cAMP partially remediates the dry mass defects of Δgpr-4 mutants on glycerol medium and Δgpr-4 strains lack the transient increase in cAMP levels observed in the wild type after addition of glucose to glycerol-grown liquid cultures. Our results support the hypothesis that GPR-4 is coupled to GNA-1 in a cAMP signaling pathway that regulates the response to carbon source in N. crassa. GPR-4-related GPCRs are present in the genomes of several filamentous ascomycete fungal pathogens, raising the possibility that a similar pathway regulates carbon sensing in these organisms.

scholarly journals Serine 129 phosphorylation of membrane-associated α-synuclein modulates dopamine transporter function in a G protein–coupled receptor kinase–dependent manner

2013 ◽  
Vol 24 (11) ◽  
pp. 1649-1660 ◽  
Author(s):  
Susumu Hara ◽  
Shigeki Arawaka ◽  
Hiroyasu Sato ◽  
Youhei Machiya ◽  
Can Cui ◽  
...  
Keyword(s):  
Cell Surface ◽  
G Protein ◽  
Dopamine Transporter ◽  
Surface Expression ◽  
Hek293 Cells ◽  
Cell Surface Expression ◽  
Receptor Kinase ◽  
Wild Type ◽  
G Protein Coupled Receptor ◽  
G Protein Coupled

Most α-synuclein (α-syn) deposited in Lewy bodies, the pathological hallmark of Parkinson disease (PD), is phosphorylated at Ser-129. However, the physiological and pathological roles of this modification are unclear. Here we investigate the effects of Ser-129 phosphorylation on dopamine (DA) uptake in dopaminergic SH-SY5Y cells expressing α-syn. Subcellular fractionation of small interfering RNA (siRNA)–treated cells shows that G protein–coupled receptor kinase 3 (GRK3), GRK5, GRK6, and casein kinase 2 (CK2) contribute to Ser-129 phosphorylation of membrane-associated α-syn, whereas cytosolic α-syn is phosphorylated exclusively by CK2. Expression of wild-type α-syn increases DA uptake, and this effect is diminished by introducing the S129A mutation into α-syn. However, wild-type and S129A α-syn equally increase the cell surface expression of dopamine transporter (DAT) in SH-SY5Y cells and nonneuronal HEK293 cells. In addition, siRNA-mediated knockdown of GRK5 or GRK6 significantly attenuates DA uptake without altering DAT cell surface expression, whereas knockdown of CK2 has no effect on uptake. Taken together, our results demonstrate that membrane-associated α-syn enhances DA uptake capacity of DAT by GRKs-mediated Ser-129 phosphorylation, suggesting that α-syn modulates intracellular DA levels with no functional redundancy in Ser-129 phosphorylation between GRKs and CK2.

Carbon source induced yeast-to-hypha transition in Candida albicans is dependent on the presence of amino acids and on the G-protein-coupled receptor Gpr1

2005 ◽  
Vol 33 (1) ◽  
pp. 291-293 ◽  
Author(s):  
M.M. Maidan ◽  
J.M. Thevelein ◽  
P. Van Dijck
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acids ◽  
Candida Albicans ◽  
Carbon Source ◽  
G Protein ◽  
High Glucose ◽  
Glucose Concentration ◽  
G Protein Coupled Receptor ◽  
Hypha Formation ◽  
G Protein Coupled

Yeast-to-hypha transition in Candida albicans can be induced by a wide variety of factors, including specific nutrients. We have started to investigate the mechanism by which some of these nutrients may be sensed. The G-protein-coupled receptor Gpr1 is required for yeast-to-hypha transition on various solid hypha-inducing media. Recently we have shown induction of Gpr1 internalization by specific amino acids, e.g. methionine. This suggests a possible role for methionine as a ligand of CaGpr1. Here we show that there is a big variation in methionine-induced hypha formation depending on the type of carbon source present in the medium. In addition high glucose concentrations repress hypha formation whereas a concentration of 0.1%, which mimics the glucose concentration present in the bloodstream, results in maximal hypha formation. Hence, it remains unclear whether Gpr1 senses sugars, as in Saccharomyces cerevisiae, or specific amino acids like methionine.

scholarly journals The Habenular Receptor GPR151 Regulates Addiction Vulnerability Across Drug Classes

2019 ◽  
Author(s):  
Beatriz Antolin-Fontes ◽  
Kun Li ◽  
Jessica L. Ables ◽  
Michael H. Riad ◽  
Andreas Görlich ◽  
...  
Keyword(s):  
G Protein ◽  
Cannabinoid Receptors ◽  
Behavioral Responses ◽  
G Protein Coupled Receptor ◽  
Inhibitory Protein ◽  
Vesicle Release ◽  
Release Probability ◽  
Drug Classes ◽  
G Protein Coupled ◽  
Camp Levels

SUMMARYThe habenula controls the addictive properties of nicotine but also densely expresses opioid and cannabinoid receptors. As such, identification of strategies to manipulate habenular activity may yield new approaches to treat substance use disorders. Here we show that GPR151, an orphan G protein-coupled receptor (GPCR) highly enriched in the habenula of humans and rodents plays a critical role in regulating habenular function and behavioral responses to addictive drugs. We show that GPR151 is expressed on axonal and presynaptic membranes and synaptic vesicles, and regulates synaptic fidelity and plasticity. We find that GPR151 associates with synaptic components controlling vesicle release and ion transport and couples to the G-alpha inhibitory protein Gαo1to reduce cAMP levels. Stable cell lines expressing GPR151 confirm that it signals via Gi/o and are amenable to ligand screens.Gpr151null mice show diminished behavioral responses to nicotine, and self-administer greater quantities of the drug, phenotypes rescued by viral re-expression ofGpr151in the habenula.Gpr151null mice are also insensitive to the behavioral actions of morphine and cannabinoids. These data identify GPR151 as a critical modulator of habenular function that controls addiction vulnerability across different drug classes.HighlightsHabenula neurons are enriched in nicotinic, opioid, cannabinoid and GPR151 receptorsGPR151 modulates synaptic fidelity and release probability at habenular terminals.Habenular GPR151 plays a role in drug abuse and food intake/weight controlGPR151 couples to the G-alpha inhibitory protein Gαo1to reduce cAMP levels.eTOC BlurbAntolin-Fontes at al. identify a G protein-coupled receptor, GPR151, which is highly enriched in human habenular neurons. These neurons are primarily enriched with nicotinic, opioid and cannabinoid receptors. We find that GPR151 modulates habenular synaptic vesicle release probability and behavioral responses to these drugs of abuse.

Transcriptional Activation of the Bradykinin Type 2 Receptor by the Tumor Suppressor P53.

Hypertension ◽  
2000 ◽  
Vol 36 (suppl_1) ◽  
pp. 719-719
Author(s):  
Samir S El-Dahr ◽  
Zubaida Saifudeen ◽  
Hong Du
Keyword(s):  
Tumor Suppressor ◽  
G Protein ◽  
Dominant Negative Mutant ◽  
Wild Type ◽  
G Protein Coupled Receptor ◽  
Water Excretion ◽  
Flanking Region ◽  
Wild Type P53 ◽  
G Protein Coupled

P145 The bradykinin type 2 receptor (BK2) is a developmentally regulated G protein-coupled receptor that mediates diverse actions such as vascular reactivity, salt and water excretion, inflammatory responses and cell growth. However, little is known regarding regulation of the BK2 gene. We report here that the rat BK2 receptor is transcriptionally regulated by the tumor suppressor protein p53. The 5’-flanking region of the rat BK2 gene contains two p53-like binding sites: a sequence at -70 bp (P1 site) that is conserved in the murine and human BK2 genes; and a sequence at -707 (P2) that is not. The P1 and P2 motifs bind specifically to p53, as assessed by gel mobility shift assays. Transient transfection into HeLa cells of a CAT reporter construct driven by 1.2-kb of the BK2 gene 5’-flanking region demonstrated that the BK2 promoter is dose-dependently activated by co-expression of wild-type p53. Co-expression of a dominant negative mutant p53 suppresses the activation of BK2 by wild-type p53. Promoter truncation localized the p53-responsive element to the region between -38 and -94 bp encompassing the p53-binding P1 sequence. Site-directed mutagenesis of the P1 site abolishes p53-mediated activation of BK2. Furthermore, p53-mediated activation of the BK2 promoter is augmented by the transcriptional co-activators, CBP/p300. Interestingly, removal of the P2 site by truncation or site-directed deletion amplifies p53-mediated activation of the BK2 promoter. These results demonstrate that the rat BK2 promoter is a target for p53-mediated activation and suggest a new physiological role for p53 in the regulation of G protein-coupled receptor gene expression.

scholarly journals A Novel High Throughput Chemiluminescent Assay for the Measurement of Cellular Cyclic Adenosine Monophosphate Levels

2000 ◽  
Vol 5 (4) ◽  
pp. 239-247 ◽  
Author(s):  
Anthony C. Chiulli ◽  
Karen Trompeter ◽  
Michelle Palmer
Keyword(s):  
G Protein ◽  
High Throughput ◽  
High Throughput Screening ◽  
Cyclic Adenosine Monophosphate ◽  
Adenosine Monophosphate ◽  
Receptor Activation ◽  
G Protein Coupled Receptor ◽  
Wide Range ◽  
G Protein Coupled ◽  
Camp Levels

The second messenger 3′, 5′-cyclic AMP (cAMP) is a highly regulated molecule that is governed by G protein-coupled receptor activation and other cellular processes. Measurement of cAMP levels in cells is widely used as an indicator of receptor function in drug discovery applications. We have developed a nonradioactive ELISA for the accurate quantitation of cAMP levels produced in cell-based assays. This novel competitive assay utilizes chemiluminescent detection that affords both a sensitivity and a dynamic assay range that have not been previously reported with any other assay methodologies. The assay has been automated in 96- and 384-well formats, providing assay data that are equivalent to, if not better than, data generated by hand. This report demonstrates the application of this novel assay technology to the functional analysis of a specific G protein-coupled receptor, neuropeptide receptor Y1, on SK-N-MC cells. Our data indicate the feasibility of utilizing this assay methodology for monitoring cAMP levels in a wide range of functional cell-based assays for high throughput screening.

scholarly journals The human cytomegalovirus G-protein coupled receptor US28 promotes latency by attenuating c-fos

2018 ◽  
Author(s):  
Benjamin A. Krishna ◽  
Monica S. Humby ◽  
William E. Miller ◽  
Christine M. O’Connor
Keyword(s):  
Host Cell ◽  
G Protein ◽  
Human Cytomegalovirus ◽  
Latent Infection ◽  
Lytic Replication ◽  
Wild Type ◽  
G Protein Coupled Receptor ◽  
In Trans ◽  
G Protein Coupled

AbstractHuman cytomegalovirus (HCMV) is a ubiquitous pathogen that undergoes latency in cells of the hematopoietic compartment, though the mechanisms underlying establishment and maintenance of latency remain elusive. We previously reported that the HCMV-encoded G-protein coupled receptor (GPCR) homolog,US28is required for successful latent infection. We now show that US28 protein (pUS28) providedin transcomplements the US28Δ lytic phenotype in myeloid cells, suggesting that sustained US28 expression is necessary for long-term latency. Furthermore, expression of pUS28 at the time of infection represses transcription from the major immediate early promoter (MIEP) within 24 hours. However, this repression is only maintained in the presence of continual pUS28 expression providedin trans. Our data also reveal that pUS28-mediated signaling attenuates both expression and phosphorylation of cellular fos (c-fos), an AP-1 transcription factor subunit, to repress MIEP-driven transcription. AP-1 binds to the MIEP and promotes lytic replication, and in line with this, we find that US28Δ infection results in an increase in AP-1 binding to the MIEP, compared to wild type latent infection. Pharmacological inhibition of c-fos represses the MIEP during US28Δ infection to similar levels we observe during wild type latent infection. Together, our data reveal that US28 is required for both establishment and long-term maintenance of HCMV latency, which is modulated, at least in part, by repressing functional AP-1 binding to the MIEP.Significance StatementHuman cytomegalovirus (HCMV) is a wise-spread pathogen that remains with an individual for life in a quiescent/latent state, posing little threat to an otherwise healthy person. However, when an individual’s immune system is severely compromised, HCMV can reactivate to its active/lytic state, resulting in viral spread and disease that is often fatal. The biological mechanisms underlying HCMV latency and reactivation remain poorly understood. Herein we show that the viral-encoded G-protein coupled receptor (GPCR)US28aids in the establishment and the maintenance of viral latency. Furthermore, we find that US28 modulates host cell proteins to suppress viral processes associated with active/lytic replication, thereby promoting latent infection. This work provides mechanism by which HCMV modulates the host cell environment to its advantage.

scholarly journals AMP010014A09 in Sus Scrofa Encodes an Analog of G Protein-Coupled Receptor 109A, Which Mediates the Anti-Inflammatory Effects of Beta-Hydroxybutyric Acid

2017 ◽  
Vol 42 (4) ◽  
pp. 1420-1430 ◽  
Author(s):  
Guangxin Chen ◽  
Shoupeng Fu ◽  
Wenqian Feng ◽  
Bingxu Huang ◽  
Shiyao Xu ◽  
...  
Keyword(s):  
G Protein ◽  
Inflammatory Cytokines ◽  
Sus Scrofa ◽  
Hydroxybutyric Acid ◽  
G Protein Coupled Receptor ◽  
Anti Inflammatory ◽  
High Level ◽  
G Protein Coupled ◽  
Camp Levels ◽  
Pro Inflammatory Cytokines

Background: Hydroxy-carboxylic acid receptor 2 (HCA2, also called GPR109A) belongs to the G protein-coupled receptor (GPCR) family and is found in humans, rats, mice, hamsters and guinea pigs, but there are almost no reports of this protein in other species. In this investigation, we speculated that AMP010014A09 (AMP+) is a homologue of GPR109A in swine. Methods: To test this hypothesis, the following experiments were designed: monocytes isolated from the peripheral blood of swine were treated with LPS after pretreating with or without β-hydroxybutyric acid (BHBA), and the levels of pro-inflammatory cytokines and inflammatory proteins were assessed. cAMP levels induced by Forskolin in swine testicular (ST) and IPEC-J2 cells were detected with or without BHBA treatment and following silencing or stable transfection of the AMP+ gene. Results: AMP+ in swine exhibited a high level of homology with HM74A in humans and PUMA-G in mice. BHBA inhibited the LPS-induced secretion of the pro-inflammatory cytokines TNF-α, IL-6 and IL-1β and the inflammatory protein COX-2 in monocytes of swine. BHBA suppressed the Forskolin-induced cAMP level increase in ST cells, but failed to inhibit the accumulation of cAMP after the AMP+ gene was silenced with shRNA by transfecting cells with the pGPU6-GFP-Neo-AMP+-sus-392 plasmid. BHBA had no effect on cAMP levels in IPEC-J2 cells, but significantly inhibited the increase in cAMP induced by Forskolin treatment following transfection of the AMP+ gene into IPEC-J2 cells by a lentivirus vector. Conclusion: Our results indicated that AMP+ encodes a G protein-coupled receptor in Sus scrofa that inhibits cAMP levels and mediates anti-inflammatory effects in swine monocytes.

Sign in / Sign up

Export Citation Format

Share Document