An endogenous A<sub>2B</sub> adenosine receptor coupled to cyclic AMP generation in human embryonic kidney (HEK 293) cells

Cooper, Joan; Hill, Stephen J.; Alexander, Stephen P.H.

doi:10.1038/sj.bjp.0701401

Cited by 53 publications

(62 citation statements)

References 16 publications

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“…3). Despite the fact that several studies had previously suggested the existence of endogenous A2b-R on HEK-293 cells (Cooper et al 1997, Gao et al 1999, Linden et al 1999, the density of this receptor appeared to be too low to contribute to calcium activity of adenosine in these cells. To our knowledge, there is no data about the expression of A3-R on HEK-293 cells.…”

Section: Endogenous Adenosine Receptorsmentioning

confidence: 88%

Adenosine does not bind to the growth hormone secretagogue receptor type-1a (GHS-R1a)

Carreira¹,

Camiña²,

Díaz‐Rodriguez³

et al. 2006

Journal of Endocrinology

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Ghrelin regulates GH secretion and energy homeostasis through the GH secretagogue receptor type-1a (GHS-R1a). This G-protein coupled receptor shows the peculiarity to transduce information provided not just by ghrelin as well as by adenosine through a supposed binding site different from the characterized ghrelin-binding pocket. Indeed, adenosine triggers intracellular calcium rise through a distinct signaling pathway to the one described for ghrelin, although it fails to stimulate GH secretion. Despite multiple active conformations of GHS-R1a, suggested as an explanation for a ligand-dependent activation of the downstream signaling, the concept of adenosine as agonist for GHS-R1a has been re-evaluated. The results revealed that calcium rise of both ghrelin and adenosine appears to be mediated by receptors that did not show the same sensitivity to protein kinase C (PKC) activity in GHS-R1a-transfected HEK 293 cells (HEK-GHS-R1a cells). The binding analyses showed the same number of adenosine-binding sites in both HEK 293 (B max Z2$01G0$15 fmol/cell) and HEK-GHS-R1a cells (B max Z1$90G0$11 fmol/cell). This binding was unaltered by different GHS-R1a antagonists. Western blot analysis showed a similar endogenous expression of endogenous adenosine receptor type-2b and -3 in both cell lines. The K d values for adenosine were 1$78 mM in HEK 293 cells and 6$30 mM in HEK-GHS-R1a cells, pointing to a modification of agonist affinity induced by overexpression of the GHSR1a. Additionally, adenosine failed to induce the GHS-R1a endocytosis, although it attenuates the ghrelin-induced GHSR1a endocytosis. In conclusion, adenosine is not an agonist of the GHS-R1a and its action is mediated by the endogenous adenosine receptor type-2b and -3, which is able to partially use the intracellular signaling machinery of HEK-GHS-R1a cells.

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Section: Endogenous Adenosine Receptorsmentioning

confidence: 88%

Adenosine does not bind to the growth hormone secretagogue receptor type-1a (GHS-R1a)

Carreira¹,

Camiña²,

Díaz‐Rodriguez³

et al. 2006

Journal of Endocrinology

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“…Also, SCH 58261 weakly antagonized both adenosine and CGS21680 responses but was ineffective against 2Cl-IB-MECA. Although VUF5574 completely antagonized the adenosine and CGS21680 dose-response functions in the double-recombinant hA3/G␣q-i3/293 cells, we cannot yet rule out the possibility that simultaneous activation of endogenous cAMP-linked A2B receptors (Cooper et al, 1997) could influence the magnitude of the A3-mediated calcium signals.…”

Section: Discussionmentioning

confidence: 99%

Pharmacological Analysis of Calcium Responses Mediated by the Human A3 Adenosine Receptor in Monocyte-Derived Dendritic Cells and Recombinant Cells

Fossetta¹,

Jackson²,

Deno³

et al. 2003

Mol Pharmacol

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Extensive characterization of adenosine receptors expressed by human monocyte-derived dendritic cells (MDDCs) was performed with quantitative polymerase chain reaction, radioligand binding, and calcium signaling. Transcript for the A3 adenosine receptor was elevated more than 100-fold in immature MDDCs compared with monocyte precursors. A3 receptor transcript was substantially diminished, and A2A receptor transcript increased, by lipopolysaccharide maturation of MDDCs. Saturation binding of N Pretreatment with 100 nM 2Cl-IB-MECA eliminated responses to CGS21680 but not to monocyte inhibitory protein-1␣. For comparison, dose-response functions were obtained from doublerecombinant human embryonic kidney 293 cells expressing the human A3 receptor and a chimeric G␣q-i3 protein, which was required to establish A3-mediated calcium signaling. The pharmacological profile of calcium signaling elicited by adenosine-related agonists in the double-recombinant cells was essentially identical to that obtained from immature MDDCs. Our results provide an extensive analysis of A3-mediated calcium signaling and unequivocally identify immature MDDCs as native expressers of the human A3 receptor.

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“…Thus, ADP exhibited only P2Y 14 -R-dependent activity at very high concentrations relative to those necessary for agonist effects of UDP. A potential explanation of the stimulatory effect of ADP in both wild-type and P2Y 14 -HEK293 cells is that ec- toenzymes convert ADP to adenosine, which in turn activates an endogenous A 2B adenosine receptor known to be present on HEK293 cells (Cooper et al, 1997;Mundell et al, 1999).…”

Section: Resultsmentioning

confidence: 99%

Quantification of G_i-Mediated Inhibition of Adenylyl Cyclase Activity Reveals That UDP Is a Potent Agonist of the Human P2Y₁₄ Receptor

Carter¹,

Fricks²,

Barrett³

et al. 2009

Mol Pharmacol

106

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The P2Y 14 receptor was initially identified as a G protein-coupled receptor activated by UDP-glucose and other nucleotide sugars. We have developed several cell lines that stably express the human P2Y 14 receptor, allowing facile examination of its coupling to native G i family G proteins and their associated downstream signaling pathways (J Pharmacol Exp Ther 330: [162][163][164][165][166][167][168] 2009). In the current study, we examined P2Y 14 receptor-dependent inhibition of cyclic AMP accumulation in human embryonic kidney (HEK) 293, C6 glioma, and Chinese hamster ovary (CHO) cells stably expressing this receptor. Not only was the human P2Y 14 receptor activated by UDP-glucose, but it also was activated by UDP. The apparent efficacies of UDP and UDP-glucose were similar, and the EC 50 values (74, 33, and 29 nM) for UDP-dependent activation of the P2Y 14 receptor in HEK293, CHO, and C6 glioma cells, respectively, were similar to the EC 50 values (323, 132, and 72 nM) observed for UDP-glucose. UDP and UDP-glucose also stimulated extracellular signal-regulated kinase (ERK) 1/2 phosphorylation in P2Y 14 receptor-expressing HEK293 cells but not in wild-type HEK293 cells. A series of analogs of UDP were potent P2Y 14 receptor agonists, but the naturally occurring nucleoside diphosphates, CDP, GDP, and ADP exhibited agonist potencies over 100-fold less than that observed with UDP. Two UDP analogs were identified that selectively activate the P2Y 14 receptor over the UDP-activated P2Y 6 receptor, and these molecules stimulated phosphorylation of ERK1/2 in differentiated human HL-60 promyeloleukemia cells, which natively express the P2Y 14 receptor but had no effect in wild-type HL-60 cells, which do not express the receptor. We conclude that UDP is an important cognate agonist of the human P2Y 14 receptor.The metabotropic P2Y receptors include a subgroup of five receptors, the P2Y 1 , P2Y 2 , P2Y 4 , P2Y 6 , and P2Y 11 receptors, that primarily signal through G q -activated signaling pathways and a subgroup of three receptors, the P2Y 12 , P2Y 13 , and P2Y 14 receptors, that primarily signal by activating heterotrimeric G proteins of the G i family (Abbracchio et al., 2006;Burnstock, 2007). The human P2Y 1 , P2Y 11 , P2Y 12 , and P2Y 13 receptors are activated by adenine nucleotides. The human P2Y 4 and P2Y 6 receptors are activated by uridine nucleotides, and the P2Y 2 receptor is activated by both ATP and UTP.

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An endogenous A_2B adenosine receptor coupled to cyclic AMP generation in human embryonic kidney (HEK 293) cells

Cited by 53 publications

References 16 publications

Adenosine does not bind to the growth hormone secretagogue receptor type-1a (GHS-R1a)

Adenosine does not bind to the growth hormone secretagogue receptor type-1a (GHS-R1a)

Pharmacological Analysis of Calcium Responses Mediated by the Human A3 Adenosine Receptor in Monocyte-Derived Dendritic Cells and Recombinant Cells

Quantification of G_i-Mediated Inhibition of Adenylyl Cyclase Activity Reveals That UDP Is a Potent Agonist of the Human P2Y₁₄ Receptor

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An endogenous A2B adenosine receptor coupled to cyclic AMP generation in human embryonic kidney (HEK 293) cells

Cited by 53 publications

References 16 publications

Adenosine does not bind to the growth hormone secretagogue receptor type-1a (GHS-R1a)

Adenosine does not bind to the growth hormone secretagogue receptor type-1a (GHS-R1a)

Pharmacological Analysis of Calcium Responses Mediated by the Human A3 Adenosine Receptor in Monocyte-Derived Dendritic Cells and Recombinant Cells

Quantification of Gi-Mediated Inhibition of Adenylyl Cyclase Activity Reveals That UDP Is a Potent Agonist of the Human P2Y14 Receptor

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An endogenous A_2B adenosine receptor coupled to cyclic AMP generation in human embryonic kidney (HEK 293) cells

Quantification of G_i-Mediated Inhibition of Adenylyl Cyclase Activity Reveals That UDP Is a Potent Agonist of the Human P2Y₁₄ Receptor