ADENOSINE-DEAMINASE AND A(1) ADENOSINE RECEPTORS INTERNALIZE TOGETHERFOLLOWING AGONIST-INDUCED RECEPTOR DESENSITIZATION

A(1) adenosine receptors (A(1)Rs) and adenosine deaminase (ADA; EC 3.5 .4.4) interact on the cell surface of DDT1MF-2 smooth muscle cells. Th e interaction facilitates ligand binding and signaling via A(1)R, but it is not known whether it has a role in homologous desensitization of A(1)Rs. Here we show that chronic exposure of DDT1MF-2 cells to the A (1)R agonist, N-6-(R)-(phenylisopropyl)adenosine (R-PIA), caused a rap id aggregation or clustering of A(1) receptor molecules on the cell me mbrane, which was enhanced by pretreatment with ADA Colocalization bet ween A(1)R and ADA occurred in the R-PLA-induced clusters. Interesting ly, colocalization between A(1)R and ADA also occurred in intracellula r vesicles after internalization of both protein molecules in response to R-PIA. Agonist-induced aggregation of A(1)Rs was mediated by phosp horylation of A(1)Rs, which was enhanced and accelerated in the presen ce of ADA Ligand-induced second-messenger desensitization of A(1)Rs wa s also accelerated in the presence of exogenous ADA, and it correlated well with receptor phosphorylation, However, although phosphorylation of A(1)R returned to its basal state within minutes, desensitization continued for hours. The loss of cell-surface binding sites (sequestra tion) induced by the agonist was time-dependent (t(1/2) = 10 +/- 1 h) and was accelerated by ADA. All of these results strongly suggest that ADA plays a key role in the regulation of A(1)Rs by accelerating liga nd-induced desensitization and internalization and provide evidence th at the two cell surface proteins internalize via the same endocytic pa thway.