Specific mutator effects of ung (uracil-DNA glycosylase) mutations in Escherichia coli.

Studies of trpA reversions revealed that G:C leads to A:T transitions were stimulated about 30-fold in E. coli ung mutants, whereas other base substitutions were not affected. A dUTPase (dut) mutation, which increases the incorporation of uracil into DNA in place of thymine, had no significant effect on the rate of G:C leads to A:T transitions. The results support the proposal that the glycosylase functions to reduce the mutation rate in wild-type cells by acting in the repair of DNA cytosine residues that have undergone spontaneous deamination to uracil. Further support was provided by the finding that when lambda bacteriophages were treated with bisulfite, an agent known to produce cytosine deamination, the frequency of clear-plaque mutants was increased an additional 20-fold by growth on an ung host. Bisulfite-induced mutations of the cellular chromosome, however, were about equal in ung+ and ung strains; it was found that during the treatment of ung+ cells with bisulfite, the glycosylase was inactivated.     

Inhibition of cyclic amp phosphodiesterase by 5'-deoxy-5'-S-isobutylthioadenosine at biologically active concentrations of drug

5'-Deoxy-5'-S-isobutylthioadenosine (SIBA), a synthetic analogue of S-adenosylhomocysteine, has been reported by others to inhibit a number of biological processes and these effects of SIBA have been attributed generally to inhibition of methyltransferases. However, the present studies with mouse lymphocytes show that SIBA also acts as a competitive inhibitor (K_i = 130 μM) of the high-affinity cyclic AMP phosphodiesterase and potentiates the cyclic AMP response of intact cells to several activators of adenylate cyclase. Moreover, SIBA has been found to inhibit lymphocyte-mediated cytolysis, a cellular function known to be sensitive to elevated lymphocyte levels of cyclic AMP, at concentrations (IC₅₀ = 250 μM) similar to those which inhibit cyclic AMP phosphodiesterase. These results indicate the need for caution in attributing biological effects of SIBA singularly to inhibition of methyltransferases and suggest the possible agency of cyclic AMP in the mechanism of SIBA action.     

Properties of Kaurene Synthetase from Marah macrocarpus Endosperm: Evidence for the Participation of Separate but Interacting Enzymes

Ent-kaurene is synthesized from geranylgeranyl pyrophosphate in a two step sequence catalyzed by kaurene synthetase; the first step (A activity) involves the conversion of geranylgeranyl pyrophosphate into the intermediate ent-trans labda-8(17), 13-dien-15-yl pyrophosphate (copalyl pyrophosphate) which is further cyclized to ent-kaurene in the second step (B activity). The resolution of enzyme fractions which catalyze each step independent of the other has been accomplished for the first time by means of QAE Sephadex A-50 chromatography and polyacrylamide gel electrophoresis of kaurene synthetase preparations from endosperm tissue of immature seed of Marah macrocarpus. Molecular weights for the A and B enzymes were each estimated as approximately 82,000 by means of gel filtration chromatography and sedimentation velocity determinations.     

Central nervous system actions of 2, 5-bis (3,4-dimethoxybenzyl) cyclopentyl amine,a peripheral dopamine blocking agent

The behavioral and brain catecholamine effects of 2,5-bis (3,4-dimethoxybenzyl) cyclopentyl amine were investigated in mice. It rapidly depleted norepinephrine. Chronic dosing also depleted dopamine, but to a lesser degree. As indicated by a lack of effect on amphetamine induced stereotypy and apomorphine induced emesis and failure to induce catalepsy, the compound does not block brain dopamine receptors. It has no effect on brain catecholamine synthesis or dopamine-β-hydroxylase activity.     

A bioenergetic study of a benthic nematode,plectus palustris de man 1880, throughout its life cycle

Summary Growth and reproduction of the parthenogenetic freshwater nematode,Plectus palustris, were studied at different controlled levels of food densities at 20° C. A bacteria-sloppy agar mixture was used as substrate and food medium. No growth or reproduction occurred at the lowest food density (8.10⁷ bacterial cells ml^-1). At 8.10⁸ cells ml^-1, the larval duration was 18.5 days, the instantaneous growth rate (g) of young larvae 0.2 d^-1 and the daily fecundity rate during a prolonged period of constant egg production 12.6 eggs·d^-1. At a food density of 8.10⁹ cells ml^-1, the corresponding values are 12.5 days, 0.4 d^-1 and 37.7 eggs d^-1.By including the data on respiration from a previous paper (Klekowski et al., 1979), the energetics of the species at different food densities can be discussed: production processes are apparently more dependent on food supply than respiration. However, prolongation of the larval phase in lower food densities greatly increases the cumulated respiratory costs per unit production. A second point is the ability to produce smaller-sized primiparous females in sub-optimal food which shortens the immature life period and serves to reduce the burden of cumulated metabolic costs for attaining sexual maturity.A comparison of the range of food densities used in the experiments with bacterial densities known from lake sediments of different trophic type suggests that food is likely to be the main factor governing the population dynamics of bacterivorous species under field situations.     

A portable pump sampler for lotic periphyton

A sampling technique for collecting lotic periphyton on sedimentary substrates using a peristaltic pump is described. Quantitative samples of periphyton standing crop and colonization rate are collected by the same procedure. The technique eliminates human disturbance problems associated with floating artificial samplers by establishing permanent sampling sites directly on submerged substances.     

GABA induces behavioral and developmental metamorphosis in planktonic molluscan larvae

Swimming planktonic larvae of the marine gastropod mollusc Haliotis rufescens require exogenous GABA or its homologs for induction of their genetically programed behavioral and developmental metamorphosis to the adult form. This requirement is stereochemically specific and absolute; GABA at 10(-6) M is fully effective in the induction of cellular differentiation, proliferation and organogenesis. The kinetics of the development of larval competence for GABA induction, and of the early metamorphic processes induced by GABA, are described. Biochemical, histological and electron micrographic analyses suggest that cyclic AMP, calcium, and a glycopeptide secretion from the cephalic sensory complex may mediate transduction of the GABA signal in the control of behavioral and morphogenetic changes induced by this environmentally deployed transmitter substance. This first observation and characterization of a major role for GABA in the control of differentiation and development, and the experimentally tractable system in which these are demonstrated, are of significance for further biomedical research.     

Measurement of surface potential and surface charge densities of sarcoplasmic reticulum membranes

Summary The binding of the anionic fluorescent probe 1-anilino-8-naphthalene-sulfonate (ANS^–) was used to estimate the surface potential of fragmented sarcoplasmic reticulum (SR) derived from rabbit skeletal muscle. The method is based on the observation that ANS^– is an obligatory anion whose equilibrium constant for binding membranes is proportional to the electrostatic function of membrane surface potential, exp(e₀/kT, where ₀ is the membrane surface potential,e is the electronic charge, andkT has its usual meaning. The potential measured is characteristic of the ANS^– bindings of phosphatidylcholine head groups and is about one-third as large as the average surface potential predicted by the Gouy-Chapman theory. At physiological ionic strength the surface potentials, measured by ANS^–, referred to as the aqueous phase bathing the surface, were in the range –10 to –15 mV. This was observed for the outside and inside surfaces of the Ca²⁺-ATPase-rich fraction of theSR and for both surfaces of theSR fraction rich in acidic Ca²⁺ binding proteins. The inside and outside surfaces were differentiated on the basis of ANS^– binding kinetics observed in stopped-flow rapid mixing experiments. A mechanism by which changes in Ca²⁺ concentration could give rise to an electrostatic potential across the membrane and possibly result in changes in Ca²⁺ permeability.The dependence of the surface potential on the monovalent ion concentration in the medium was used together with the Gouy-Chapman theory to determine the lower limits for the surface charge density for the inside and outside surfaces of the two types ofSR. Values for the Ca²⁺-ATPase richSR fraction were between 2.9×10³ and 3.8×10³ esu/cm², (0.96×10^–6 and 1.26×10^–6 C/cm²) with no appreciable transmembrane asymmetry. A small amount of asymmetry was observed in the values for the inside and outside surfaces of the fraction rich in acidic binding proteins which were ca. 6.6×10³ and ca. 2.2×10³ esu/cm² (2.2×10^–6 and 0.73×10^–6 C/cm). The values could be accounted for by the known composition of negatively-charged phospholipids in theSR. The acidic Ca²⁺ binding proteins were shown to make at most a small contribution to the surface charge, indicating that their charge must be located at least several tens of Å from the membrane surface. The experiments gave evidence for a Donnan effect on the K⁺ distribution in the fraction rich in acidic binding proteins. This could be accounted for by the known concentration of acidic binding proteins in thisSR fraction.The equilibrium constant for ANS^– was shown to be more sensitive to changes in the divalent cation concentration than to changes in the monovalent cation concentration, as predicted by the Gouy-Chapman theory. Use of these findings together with the stopped-flow rapid mixing techniques constitutes a method for rapid and continuous monitoring of changes in ion concentrations in theSR lumen.     

Metabolic formation of nucleoside-modified analogues of S-adenosylmethionine

A Pseudo-ovalbumin gene, bearing significant nucleotide sequence homology to the ovalbumin gene, has been cloned from genomic chick DNA. Similar to the authentic ovalbumin gene, the pseudo-gene is a unique sequence gene in the chick genome and is expressed at a low level in the immature chick oviduct. In contrast to the ovalbumin gene, expression of the pseudo-gene in the oviduct is not inducible by estrogen. The concentration of pseudo-gene RNA is only ～0.01% of that of authentic ovalbumin mRNA in estrogen-stimulated oviduct cells. Nucleotide sequence analysis of the two sequence related genes may reveal the molecular basis of differential response to steroid hormone induction in the same tissue.     

Purine transport by Malpighian tubules of pteridine-deficient eye color mutants of Drosophila melanogaster

Uptakes of guanine into Malpighian tubules of wild-type Drosophila and the eye color mutants white (w), brown (bw), and pink-peach (p ^p) have been compared. Tubules for each of these mutants are unable to concentrate guanine intracellularly. The transport of xanthine and riboflavin is also deficient in w tubules. The transport of guanosine, adenine, hypoxanthine, and guanosine monophosphate is similar in wild-type and white Malpighian tubules. These data and other information about these mutants make it likely that these pteridine-deficient eye color mutants do not produce pigments because of the inability to transport a pteridine precursor. This view supports the hypothesis that mutants which lack both pteridine and ommochromes do so because precursors to both classes of pigments share a common transport system.This work was supported by Grant GM22366 from NIH.