TISSUE-SPECIFIC EFFECTS OF DIVALENT-CATIONS AND ACTIVATORS ON SOLUBLE GUANYLATE CYCLASE

The interactions of divalent cations, Mn2+, Mg2+, and Ca2+, with the cytosolic guanylate cyclase (GTP pyrophosphate-lyase (cyclizing, EC 4.6.1.2) from different tissues were studied. Guanylate cyclase activities of the kidney, liver, and lung were strongly dependent on Mn2+. In contrast, the enzyme in smooth muscle of the colon, aorta, and vas diferens was active with Mg2+ as well as with Mn2+. Ca2+ was ineffective in all tissues. Preincubation, at 30°C, of colon extracts, but not those of kidney and liver, increased guanylate cyclase activity. The Mg2+-dependent activity was preferentially enhanced by this treatment. These results suggest that when the enzyme was autoactivated by endogenus factors it became more Mg2+ dependent. Dithiothreitol strongly inhibited the Mg2+-dependent colon enzyme, whereas activities in kidney and liver were not affected and the response of the enzyme in lung was intermediate. This suggests that autoactivation involved an oxidative-reductive alteration of the enzyme. Ca2+ markedly inhibited the Mg2+-dependent activity in smooth muscles but Mg2+-dependent activities in lung, liver, and kidney were not influenced appreciably. Exogenous activators, dehydroascorbate and NaN3, increased guanylate cyclase, assayed with either Mg2+ or Mn2+. However, the relative stimulation of the enzyme assayed with Mg2+ was greater than with Mn2+. When activated by these exogenous agents, guanylate cyclase in all tissues became inhibitable by Ca2+. These findings suggest that guanylate cyclase in smooth muscle, as prepared, was in a partially activated form. The endogenous activating factors in colon smooth muscle were heat-stable, largely extractable with chloroform/methanol, and cochromatographed with authentic fatty acids. Arachidonic acid stimulated colon guanylate cyclase and enhancement of the Mg2+-dependent activity was blocked by Ca2+. This strongly infers that a significant part of the endogenous activating factors in the colon was fatty acids or their derivatives. The colon activators had only minimal affects on the enzyme in the kidney. Similarly prepared activator extracts from the kidney increased colon guanylate cyclase but did not stimulate the renal enzyme. Thus, the ability of the enzyme to be stimulated by endogenous activators was dependent on the tissue from which the enzyme was derived. The possibility that the interactions of Mg2+ and Ca2+ on guanylate cyclase in smooth muscles are of regulatory significance to the contractile response of the muscle was discussed. It was proposed as a working hypothesis that cyclic GMP and Ca2+ might participate in reciprocal negative feedback mechanisms. © 1979.