Adenosine Deaminase and A1 Adenosine Receptors Internalize Together following Agonist-induced Receptor Desensitization

Abstract

Adenosine is an autacoid that exerts its physiological actions through specific cell surface receptors. Four adenosine receptors (A 1 , A 2A , A 2B , and A 3 ), belonging to the family of G protein-coupled receptors, have been cloned and pharmacologically characterized (1). Acting through different adenosine receptors, adenosine is a neuromodulator in both the central and peripheral nervous systems (2, 3). Via A 1 adenosine receptors (A 1 Rs), 1 adenosine reduces heart rate (4), glomerular filtration rate, and renin release in the kidney (5); it induces bronchoconstriction (6, 7) and inhibits lipolysis. A 1 Rs can be coupled to different pertussis toxin-sensitive G proteins (8 -10), which mediate inhibition of adenylate cyclase (11) and regulate Ca 2ϩ and K ϩ channels and inositol phosphate metabolism (1). A 1 R displays two different affinities for agonist, which have classically been attributed to a different coupling to heterotrimeric G proteins (12). According to this two-independent site model, coupled receptor-G protein complexes display high affinity for agonists (K d ϭ 0.1-0.2 nM), and uncoupled receptors display low affinity (1-2 nM) (12, 13). The recently reported cluster-arranged cooperative model predicts that the high and low affinity sites are a consequence of the negative cooperativity of agonist binding and do not seem to be related to the content of free and G protein-coupled receptors (14). According to the cluster-arranged cooperative model, the agonist-induced conversion of the high to the low affinity state may partially explain the ligand-induced desensitization of A 1 R (14, 15).Receptors belonging to the G protein-coupled receptor family are desensitized and down-regulated in response to agonist stimulation. ␤ 2 -Adrenergic receptors (␤ 2 -ARs) constitute the best characterized system within the G protein-coupled receptor family. Agonist-induced ␤ 2 -AR desensitization is caused by a conformational change of the agonist-occupied receptor that facilitates receptor phosphorylation by second messenger-activated kinases or G protein-coupled receptor kinases (16,17). Following ␤ 2 -AR phosphorylation, ␤-arrestin binds to the phosphorylated receptor and uncouples the receptor from the heterotrimeric G proteins (18,19). ␤-Arrestin not only desensitizes the receptor but also functions as a clathrin adaptor, mediating receptor sequestration, i.e. receptor internalization toward intracellular compartments (20,21). Although sequestration is not required for phosphorylation and desensitization, it appears to be necessary for dephosphorylation and resensitization of ␤ 2 -ARs (22, 23).Like other G protein-coupled receptor members, A 1 R expression is regulated in response to agonist or antagonist stimulation. Desensitization of A 1 R has been described in intact animals and in cell cultures. Prolonged administration of A 1 R agonists to animals leads to functional desensitization of A 1 R in guinea pig heart (24), rat adipocytes (25), rat atrial muscle (26), and rat brain (27,28). The reduced functiona...

The Heat Shock Cognate Protein hsc73 Assembles with A1 Adenosine Receptors To Form Functional Modules in the Cell Membrane

“…R-PIA-induced internalization of A 1 Rs results in sequestration of the receptor in intracellular vesicles, an effect that is potentiated in the presence of ADA (3,28). Internalization of A 1 R was monitored by confocal microscopy after culturing the cells in the presence of a fluorochrome-conjugated anti-A 1 R antibody (PC21), ADA, and the ligand R-PIA (see reference 3).…”

“…On the other side of the plasma membrane, A 1 Rs interact with ecto-adenosine deaminase (ecto-ADA) an interaction which is needed for efficient signaling (11,27). Moreover, in response to agonists, A 1 Rs and ADA are internalized together via the same endocytic pathway (28).…”

Adenosine A1 receptor agonist N6-cyclohexyl-adenosine induced phosphorylation of delta opioid receptor and desensitization of its signaling

“…Previous studies showed that A 1 R agonist (R)-PIA induced a time-dependent reduction in cell surface adenosine A1 receptor radioligand binding sites, which reached a maximum at 48-72 h [31,32] . To examine whether heterologous desensitization of the DOR by CHA was attributed to receptor down-regulation, saturation binding was used to assess receptor affinity (K d ) and receptor density (B max ) of DOR in plasma membranes prepared from cells pretreated with or without 1 μmol/L CHA for 72 h. Saturation curves of DOR ( Figure 5A) and the Scatchard analysis of the saturation binding ( Figure 5B) were present in Figure 5, which showed no significant change of receptor numbers and affinity after chronic CHA exposure.…”

Comodulation of CXCR4 and CD26 in Human Lymphocytes

“…In fact, the lack of the cytoplasmic tail, which is known to prevent SDF-1␣Ϫinduced down-regulation of CXCR4, also prevents internalization of CD26. These results indicate that treatment of lymphocytes with chemokines or agents that activate protein kinase C leads to the simultaneous internalization of CXCR4 and CD26, probably via the same endocytic pathway, as we have demonstrated for other interacting cell surface proteins (35). SDF-1␣-induced internalization of CXCR4 is mediated by coated vesicles (17).…”

“…This interesting finding is evidence for a G i protein-independent signaling pathway that regulates the SDF-1␣-induced simultaneous internalization of the two proteins. To our knowledge this is the second example of ligand-induced cointernalization of a receptor and the enzyme that inactivates the ligand for the receptor (35). In these models of interacting proteins, the existence of membrane-bound and soluble forms of degrading enzymes is relevant.…”