Adenosine cyclic 3',5'-monophosphate uptake and regulation of membrane protein kinase in intact human erythrocytes

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Abstract
The uptake of adenosine cyclic[c]AMP and stimulation of membrane-associated protein kinase in mature human erythrocytes were investigated. cAMP transport across the membrane was temperature-dependent; cAMP binding to the isolated membrane had less temperature dependence. More than 99% of the [3H] cAMP taken up by erythrocytes was nonmembrane bound. Maximal stimulation of membrane protein kinase and maximal occupancy of membrane cAMP binding sites by extracellular cAMP occurred at 30.degree. C within 30 min after initiation of the incubation of erythrocytes with cAMP. The concentration of extracellular cAMP that gave half-maximal stimulation of membrane protein kinase was 5.4 .times. 10-4 M, a value consistent with the concentrations of cAMP (5.2 .times. 10-4 M) found to occupy half-maximally the membrane cAMP binding sites in erythrocytes. Extracellular cAMP, and to a lesser extent cyclic GMP and cyclic inosine 3'',5''-monophosphate, stimulated membrane protein kinase in erythrocytes. The cAMP uptake by human erythrocytes and cAMP binding to erythrocyte membranes were blocked by an inhibitor [4,4''-bis(isothiocyano)stilbene-2,2-disulfonate] on the anion channel. cAMP can be transported across membranes into human erythrocytes and can bind to membranes to activate membrane protein kinase. It appears that there is a shared transport channel for cAMP and anion transport.