Loss of the PGE1 requirement for MDCK cell growth associated with a defect in cyclic AMP phosphodiesterase

Prostaglandin E₁(PGE₁), one of the components in the hormone-supplemented, serum-free medium for Madin Darby Canine Kidney (MDCK) cells (Medium K-1), is required for both long-term growth and for dome formation. Variant cells have been isolated from MDCK populations, which lack the PGE₁, requirement for long-term growth in Medium K-1. These variants will be useful in identifying the molecular events initiated by PGE₁ which are necessary for the growth response to be observed. The growth and functional properties of five independently isolated PGE₁ independent clones have been examined. Normal MDCK cells grew at an equivalent rate in Medium K-1 and in serum-supplemented medium; the growth rate was lower in Medium K-1 lacking PGE₁. In contrast, PGE₁ independent clone 1 grew at an equivalent rate in Medium K-1 minus PGE₁, and in serum-supplemented medium. When PGE₁ was added to K-1 minus PGE₁, less growth of PGE₁ independent clone 1 was observed. A similar observation was made with one other PGE₁ independent clone which was studied. A hormone deletion study indicated that PGE₁ independent clone 1 still retained growth responses to the other four supplements in Medium K-1 (insulin, transferrin, T₃, and hydrocortisone). The molecular alterations associated with loss of the PGE₁ requirement for long-term growth were examined. At confluency, all of the PGE₁ independent clones studied had higher intracellular cyclic AMP levels following PGE₁ treatment, as compared with normal MDCK cells. The increased cyclic AMP levels in the variant cells could result from a number of different types of defects, including reduced cyclic adenylic acid (cyclic AMP) efflux, an increased affinity of PGE₂ for the PGE₁ receptor, or a defect in cyclic AMP metabolism. However, in all of the variant clones studied a decreased rate of cyclic AMP degradation by cyclic AMP phosphodiesterase was observed. Thus, the increased cyclic AMP levels in the PGE₁ independent variants may result from alterations which affect cyclic AMP metabolism. The effect of PGE₁ on dome formation by the variant cells was also examined. The frequency of dome formation by PGE₁ independent clone 1 was enhanced in a dosage-dependent manner, like normal MDCK cells. This observation suggests that PGE₁ affects MDCK cell growth and dome formation by different mechanisms.     

Effects of Kinetin, IAA, and Gibberellin on Ethylene Production, and Their Interactions in Growth of Seedlings

Kinetin in concentrations of 10⁻⁸ to 10⁻⁴ m, stimulated ethylene production in 3 and 4-day old etiolated seedlings of Alaska pea (Pisum sativum L. var. Alaska). Seedlings of other species responded similarly. The response to kinetin depended on the age of the seedlings.     

Derepression of Phosphomannose Isomerase by Regulator Gene Mutations Involved in Capsular Polysaccharide Synthesis in Escherichia coli K-12

A regulator gene mutation (capR) that causes increased synthesis of capsular polysaccharide and derepressed synthesis of several enzymes involved in polysaccharide synthesis also derepresses phosphomannose isomerase (PMI) synthesis. In contrast, a second mutation (capS, which maps separately from capR) that causes increased production of the same polysaccharide does not lead to increased synthesis of PMI (nor of several of the other enzymes involved in polysaccharide synthesis). Introduction of the capR9 allele by transduction or mutation of capR(+) to capR can change the phenotype of a mannose-negative nonmucoid strain to a mannose-positive mucoid phenotype. Thus, genotype capR(+)man-2 is mannose-negative and nonmucoid, but genotype capR9 man-2 is mannose positive and mucoid. Other interactions between these alleles in the synthesis of capsular polysaccharide are recorded.     

Physiological Control of Molluscan Gill Cilia by 5-Hydroxytryptamine

An examination is made of the hypothesis that endogenous 5-hydroxytryptamine (5-HT) serves as a local hormone regulating ciliary activity in the lamellibranch gill. These cilia are sensitive to exogenous 5-HT and respond to it by a prompt, sustained, and reversible rise in beat frequency; at the same time the carbohydrate metabolism is stimulated, as described elsewhere. Control gill contains small but definite amounts of endogenous 5-HT according to bioassay, fluorometry, and chromatography. The amount can be increased markedly by exposing the isolated gill to the precursor substance 5-hydroxytryptophan but not l-tryptophan. As the tissue level of 5-HT rises, the spontaneous beat frequency also rises. Both remain elevated for hours and perhaps for days. The gill of Mytilus edulis is richer than the gill of Modiolus demissus in both endogenous 5-HT and effective 5-hydroxytryptophan decarboxylase activity. Modiolus gill lacks the 5-hydroxyindole oxidase by which Mytilus gill destroys 5-HT. What if any mechanism exists in Modiolus for degrading 5-HT is not known, but monoamine oxidase is not present. The 5-HT content of Mytilus and Modiolus gill cannot be modified by treatment with reserpine or α-methyl-dopa. Which cells of the gill synthesize and destroy 5-HT has not been established, but these observations support the concept that the physiological activity of lamellibranch gill cilia is controlled by a serotonergic mechanism.