Competitive Inhibition of Beef Heart Cyclic AMP Phosphodiesterase by Cytokinins and Related Compounds

Abstract

Two cytokinins and four related analogs, none of which is a cyclic ribonucleotide, have been shown to act as competitive inhibitors of the high Km cyclic-AMP phosphodiesterase (3': 5'-cyclic-AMP 5'-nucleotidohydrolase, EC 3.1.4.1-7) activity from beef heart. Weak inhibition of the low Km cyclic AMP phosphodiesterase activity was also observed, suggesting a possible mechanism for regulation of intracellular cyclic AMP levels by the exogenously added compounds. In addition to the kinetic data, obtained on the six inhibitors in four different heterocyclic series, 15 other cytokinins and related compounds have been shown to inhibit the high Km cyclic AMP phosphodiesterase activity at single concentrations of substrate and inhibitor. Heterocycles such as adenosine and 7-amino-3-methiylpyrazolo [4,3-d]pyrimidine, which lack the N-substituent, were inactive as cyclic AMP phosphodiesterase inhibitors. The observed inhibition of cyclic AMP phoplhodiesterase supports prior observations which implicate exogenously added cytokinins in cyclic AMP metabolism.Cytokinins were first isolated as plant factors responsible for the promotion of cell division and growth (1, 2). The compounds, which are typically N6-substituted adenine and adenosine derivatives, occur at the purine, ribonucleoside, and ribonucleotide levels in plants, as well as in the transfer RNAs of most forms of life (2-5). In spite of the widespread natural occurrence of cytokinins and the diverse metabolic effects which they are now known to promote in both plants and animals (1-13), relatively little has been learned about their mechanism of action at the molecular level. This prompted the design and synthesis of a class of potent anticytokinins, structurally related to the cytokinins, in the hope that the antimetabolites might extend the study of cytokinins to new biological systems and provide useful information pertinent to the mechanism of cytokinin action (14)(15)(16)(17).On the basis of the similar effects obtained with N6-(A2-isopentenyl)adenosine and dibutyryl cyclic AMP, the cytokinins have been postulated to mediate their effects in phytohemagglutinin-treated human lymphocytes by involvement in cyclic AMP metabolism (13). This postulate was consistent with the finding that the cytokinin trans-zeatin ribonucleoside was inhibitory to the cyclic AMP phosphodiesterases from beef brain and crown-gall tumor cells, the latter of which was Abbreviations: Dibutyryl cyclic AM1P, N6,02'-(dibutyryl)adenosine cyclic 3': 5'-monophosphate; cyclic AMP, adenosine cyclic 3': 5'-monophosphate; trans-zeatin ribonucleoside, N6-(4-hydroxy-3-methyl-trans-2-butenyl)adenosine; 8-bromo

Ligand Specificity of a High Affinity Cytokinin-binding Protein

“…The specificity of this set of interactions with chloroplast electron transport inhibitors is arguable since structurally disparate energy transfer inhibitors (DCCD and organotin compounds) are also CBP ligands. Certain cytokinins have been shown to inhibit competitively mammalian 3',5'-cyclic AMP phosphodiesterase (12), and MIX, papaverine, 2',3'-cyclic AMP, and alkylated 3',5'-cyclic AMP derivatives (all CBP ligands) also bind to this enzyme (1). The Kd of CBP for MIX (5 x 10-M) is similar to that of beef heart 3',5'-cycic AMP phosphodiesterase for this compound (2), MIX being one of the more effective inhibitors of the enzyme.…”

Characterization of Cyclic Nucleotide Phosphodiesterase Activity in Sunflower Callus

“…A recent report on cytokinins (2 m M ) being potent inhibitors of 3′, 5′‐cAMP phosphodiesterase from beef heart (15) led us to do a similar investigation on the sunflower enzyme. The compounds tested were 6( γ,γ ‐dimethyl‐allylamino)‐purine, kinetin (6‐furfurylamino‐purine), kinetin‐riboside (6‐furfuryl‐amino‐purine riboside), zeatin (6‐{trans‐4‐hydroxy‐3‐methylbut‐2‐enylamino}purine), and zeatin riboside.…”

Anticytokinin activity of substituted pyrrolo[2,3- d ]pyrimidines

“…To 400 mg (2.0 mmol) of 4-chloro-2-methylthiopyrrolo [2,3-d]pyrimidine (18) 4-Cyclopentylamino-6-methylthiopyrrolo[2,3-dlpyrimidine-5-carbonitrile (11). To 533 mg (2.61 mmol) of the presumed methoxymethylenamino derivative of 2-amino-3,4-dicyano-5-thiopyrrole (20) was added 3 ml of cyclopentylamine and the mixture was heated at reflux for 2 hr under N2.…”

“…N6-(3-Methyl-2-butenyl)adenosine (i6Ado) has been reported to be an immunosuppressive agent (7) and to induce hematological remissions in leukemia patients (8,9). Several cytokinins, including i6Ado, have also been shown to be capable of regulating the growth of phytohemagglutinin-transformed human lymphocytes (10 and references therein), presumably by involvement in cyclic AMP metabolism, and i6Ado was shown to act as a competitive inhibitor of beef heart cyclic AMP phosphodiesterase (11).…”