Adenosine is emerging as a key modulator of metabolism. It acts both as an intracellular and intercellular messenger. The existence of extracellular plasma membrane receptors for adenosine is well documented. At least two types of such receptors exist; one activates and the other depresses the activity of adenyl cyclase. Other types of plasma membrane adenosine receptors may exist. Intracellular adenosine receptors are not well documented; binding of adenosine to these receptors results in a decrease of cAMP, but effects not involving cAMP are not ruled out. We have isolated several adenosine receptors in pure form by affinity chromatography of cell membranes and cytosol from heart tissue and aortic smooth muscle. These proteins are not any of the known enzymes of adenosine metabolism nor are they proteolytic artifacts of these enzymes. They are eluted from affinity columns specifically by adenosine and some of its analogs, but not by cAMP or AMP. We will characterize these proteins and study their function. We will investigate whether one or more of the proteins that we have isolated act as modulators of the activity of other enzymes. We will study whether our adenosine-binding proteins specifically interact with other proteins, much like the regulatory subunits of cyclic AMP-dependent protein kinases bind to the catalytic subunits, or like calmodulin binds to some phosphodiesterases, or the GTP-modulator protein binds to adenyl cyclase. We have observed that adenosine inhibits the phosphorylation of myosin light chain in extracts of smooth muscle. The adenosine receptor involved in this inhibition will be investigated. Smooth muscle possesses a different spectrum of adenosine receptor proteins compared to heart and liver, one of these may be involved in modulating myosin light chain phosphorylation. We have partially purified a soluble 5'-nucleotidase from heart. We will study the regulatory properties of this enzyme and purify it to homogeneity. We have partially purified a plasma membrane ATP pyrophosphatase from heart. It is proposed to purify this enzyme to homogeneity and to study its properties with the aim of understanding its function.
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