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In Situ Reconstitution of the Adenosine A2A Receptor in Spontaneously Formed Synthetic Liposomes.

ABSTRACT: Cell transmembrane receptors play a key role in the detection of environmental stimuli and control of intracellular communication. G protein-coupled receptors constitute the largest transmembrane protein family involved in cell signaling. However, current methods for their functional reconstitution in biomimetic membranes remain both challenging and limited in scope. Herein, we describe the spontaneous reconstitution of adenosine A2A receptor (A2AR) during the de novo formation of synthetic liposomes via native chemical ligation. The approach takes advantage of a nonenzymatic and chemoselective method to rapidly generate A2AR embedded phospholiposomes from receptor solubilized in n-dodecyl-?-d-maltoside analogs. In situ lipid synthesis for protein reconstitution technology proceeds in the absence of dialysis and/or detergent absorbents, and A2AR assimilation into synthetic liposomes can be visualized by microscopy and probed by radio-ligand binding.

SUBMITTER: Brea RJ

PROVIDER: S-EPMC5702554 | biostudies-literature | 2017 Mar

REPOSITORIES: biostudies-literature

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In Situ Reconstitution of the Adenosine A2A Receptor in Spontaneously Formed Synthetic Liposomes.

Brea Roberto J RJ Cole Christian M CM Lyda Brent R BR Ye Libin L Prosser R Scott RS Sunahara Roger K RK Devaraj Neal K NK

Journal of the American Chemical Society 20170306 10

Cell transmembrane receptors play a key role in the detection of environmental stimuli and control of intracellular communication. G protein-coupled receptors constitute the largest transmembrane protein family involved in cell signaling. However, current methods for their functional reconstitution in biomimetic membranes remain both challenging and limited in scope. Herein, we describe the spontaneous reconstitution of adenosine A2A receptor (A2AR) during the de novo forma ...[more]

PMID: 28263576

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Project description:ObjectivePharmacological activation of adenosine signaling has been shown to increase ?-cell proliferation and thereby ?-cell regeneration in zebrafish and rodent models of diabetes. However, whether adenosine has an endogenous role in regulating ?-cell proliferation is unknown. The objective of this study was to determine whether endogenous adenosine regulates ?-cell proliferation-either in the basal state or states of increased demand for insulin-and to delineate the mechanisms involved.MethodsWe analyzed the effect of pharmacological adenosine agonists on ?-cell proliferation in in vitro cultures of mouse islets and in zebrafish models with ?- or ?-cell ablation. In addition, we performed physiological and histological characterization of wild-type mice and mutant mice with pancreas- or ?-cell-specific deficiency in Adora2a (the gene encoding adenosine receptor A2a). The mutant mice were used for in vivo studies on the role of adenosine in the basal state and during pregnancy (a state of increased demand for insulin), as well as for in vitro studies of cultured islets.ResultsPharmacological adenosine signaling in zebrafish had a stronger effect on ?-cell proliferation during ?-cell regeneration than in the basal state, an effect that was independent of the apoptotic microenvironment of the regeneration model. In mice, deficiency in Adora2a impaired glucose control and diminished compensatory ?-cell proliferation during pregnancy but did not have any overt phenotype in the basal state. Islets isolated from Adora2a-deficient mice had a reduced baseline level of ?-cell proliferation in vitro, consistent with our finding that UK432097, an A2a-specific agonist, promotes the proliferation of mouse ?-cells in vitro.ConclusionsThis is the first study linking endogenously produced adenosine to ?-cell proliferation. Moreover, we show that adenosine signaling via the A2a receptor has an important role in compensatory ?-cell proliferation, a feature that could be harnessed pharmacologically for ?-cell expansion and future therapeutic development for diabetes.

Dataset Information

In Situ Reconstitution of the Adenosine A2A Receptor in Spontaneously Formed Synthetic Liposomes.

Publications

In Situ Reconstitution of the Adenosine A<sub>2A</sub> Receptor in Spontaneously Formed Synthetic Liposomes.

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