Relationship between Legionella pneumophila and Acanthamoeba polyphaga: physiological status and susceptibility to chemical inactivation.

Survival studies were conducted on Legionella pneumophila cells that had been grown intracellularly in Acanthamoeba polyphaga and then exposed to polyhexamethylene biguanide (PHMB), benzisothiazolone (BIT), and 5-chloro-N-methylisothiazolone (CMIT). Susceptibilities were also determined for L. pneumophila grown under iron-sufficient and iron-depleted conditions. BIT was relatively ineffective against cells grown under iron depletion; in contrast, iron-depleted conditions increased the susceptibilities of cells to PHMB and CMIT. The activities of all three biocides were greatly reduced against L. pneumophila grown in amoebae. PHMB (1 x MIC) gave 99.99% reductions in viability for cultures grown in broth within 6 h and no detectable survivors at 24 h but only 90 and 99.9% killing at 6 h and 24 h, respectively, for cells grown in amoebae. The antimicrobial properties of the three biocides against A. polyphaga were also determined. The majority of amoebae recovered from BIT treatment, but few, if any, survived CMIT treatment or exposure to PHMB. This study not only shows the profound effect that intra-amoebal growth has on the physiological status and antimicrobial susceptibility of L. pneumophila but also reveals PHMB to be a potential biocide for effective water treatment. In this respect, PHMB has significant activity, below its recommended use concentrations, against both the host amoeba and L. pneumophila.     

Early gene interaction during prepupal expression of Drosophila arginine kinase.

Arginine kinase displays a distinctive rise and fall in specific activity and specific protein levels during the prepupal stage of Drosophila development with maximal activity occurring at morphological stage P3. This developmentally regulated peak is under the influence of ecdysone. Altered doses of the major ecdysone-inducible "early" genes at cytological regions 75B and 2B5 alter this pattern of expression while altered doses of another major "early" gene at 74EF have no effect. We hypothesize that a product of the 2B5 locus and a product of the 75B locus interact to effect this developmental pattern of expression of Drosophila arginine kinase.     

Anthrax toxin protective antigen: low-pH-induced hydrophobicity and channel formation in liposomes

To probe the role of the protective antigen (PA) component of anthrax toxin in toxin entry into animals cells, we examined the membrane channel-forming properties and hydrophobicity of intact and trypsin-cleaved forms of the protein at various pH values. At neutral pH neither form caused release of entrapped K+ from unilamellar lipid vesicles. At pH values below 6.0, however, K+ was rapidly released upon addition of either the nicked PA (PAN) or the 63 kDa tryptic fragment of PA (PA63), which has been implicated in the toxin entry process. Under the same conditions intact PA exhibited only weak channel-forming activity, and PA20, the complementary tryptic fragment, showed no such activity. Both PA and PA63 exhibited enhanced hydrophobicity at acidic pH values, but the enhancement was greater and the pH threshold higher with PA63. Our findings indicate that proteolytic removal of PA20 from intact PA enables the residual protein, PA63, to adopt a conformation at mildly acidic pH values that permits it to insert readily and form channels in membranes. Thus acidic conditions within endocytic vesicles may trigger membrane insertion of PA63, which in turn promotes translocation of ligated effector moieties, edema factor or lethal factor, across the vesicle membrane into the cytosol.     

Early gene interaction during prepupal expression of Drosophila arginine kinase

Arginine kinase displays a distinctive rise and fall in specific activity and specific protein levels during the prepupal stage of Drosophila development with maximal activity occurring at morphological stage P3. This developmentally regulated peak is under the influence of ecdysone. Altered doses of the major ecdysone-inducible “early” genes at cytological regions 75B and 2B5 alter this pattern of expression while altered doses of another major “early” gene at 74EF have no effect. We hypothesize that a product of the 2B5 locus and a product of the 75B locus interact to effect this developmental pattern of expression of Drosophila arginine kinase. © 1992 Wiley-Liss, Inc.     

Effect of length, supercoiling, and pH on intramolecular triplex formation. Multiple conformers at pur.pyr mirror repeats

The conformations adopted by five oligopurine.oligopyrimidine (pur.pyr) inserts of various lengths and sequence repeats in recombinant plasmids were evaluated as a function of pH and negative super-helicaldensity. Patterns of chemical reactivity (OsO4 and diethylpyrocarbonate) indicate that long (greater than 36 base pairs) pur.pyr segments can adopt intramolecular triplexes and that increasing the length of the pur.pyr tract reduces the dependence on low pH for structure formation, such that (GA)37 adopts an intramolecular triplex under moderate levels of negative superhelical stress (-sigma = 0.049) at neutral pH. This demonstrates that long pur.pyr segments, which are abundant in eukaryotic genomes, have the potential to adopt triplexes in vivo. Two-dimensional gel electrophoresis of the plasmids combined with chemical probing indicates that for longer sequences, multiple conformers of the intramolecular triplex exist at low pH. These conformers result from nucleation at various positions on the polypurine stretch, giving rise to different extents of relaxation at the same linking number. In addition, the metal ions Co2+, Mn2+, and Mg2+ have profound effects on the pattern of chemical reactivity displayed by long pur.pyr segments at both neutral and low pH, indicating that quite different structures may form in the presence of divalent metal ions. Thus, the types and extent of unusual structures adopted by long pur.pyr segments are complex and heterogeneous, and are dependent on pH, supercoiling, and the presence of divalent cations.     

Isolation and genetic characterization of streptomycin and chloramphenicol resistant mutants in the mossPhyscomitrella patens

Summary Mutant strains of the mossPhyscomitrella patens that were resistant to the antibiotics streptomycin and chloramphenicol were isolated following UV irradiation. Mutants resistant to streptomycin at 1.7×10⁻⁴ M showed both dominant and recessive Mendelian and inheritance patterns. Mutants resistant to streptomycin at 1.7 ￠ 10⁻⁴ M and to chloramphenicol at 3.1×10⁻⁴ M were, with one exception, cytoplasmically inherited.     

2-Decarboxy-2-hydroxymethyl prostaglandin E1 (TR4161), a prostaglandin bronchodilator of low tracheobronchial irritancy

2-Decarboxy 2-hydroxymethyl prostaglandin E₁ (TR4161) relaxed isolated guinea-pig trachea with about double and relaxed human isolated bronchial muscle with about one half the potency of PGE₁. In conscious restrained cats an aerosol of TR4161 was about 100–1000 times less active than PGE₁ in inducing tracheobronchial irritation. When given intravenously or by aerosol to the anaesthetised spontaneously breathing guinea-pig, TR4161 was approximately equipotent with PGE₁ in inhibiting histamine-induced bronchoconstriction and in reducing basal inherent tone. The onset and duration of the bronchodilator effects of TR4161 administered intravenously, however, were significantly longer than those of PGE₁. In conscious guinea-pigs, TR4161 by aerosol was approximately three times more potent than PGE₁ in preventing histamine-induced convulsions, whereas only TR4161 was active in this test system when the test drugs were administered orally. These observations indicate that TR4161 might be therapeutically useful as a non-irritant prostaglandin bronchodilator in conditions of airway obstruction.     

Inhibition of prostaglandin biosynthesis by sulphasalazine and its metabolites

Sulphasalazine (SZ) inhibits prostaglandin (PG) biosynthesis $\text{in vitro}$ with a potency comparable to that of aceylsalicylate. The metabolites of SZ, sulphapyridine and 5-aminosalicylic acid, were of considerably lower potency as inhibitors of PG biosynthesis in the synthetase preparations used. The inhibition of prostaglandin production by SZ could at least partly account for the clinical utility of sulphasalazine in ulcerative colitis. Sulphapyridine may help to maintain inhibitory concentrations of SZ by restraining bacterial breakdown of the active drug.     

Prostaglandins, cyclic AMP and the mechanism of opiate dependence.

Further evidence is given that dependence arises from the agonist action of opiates. From this and our previous propositions assigning a fundamental role to neuronal cyclic AMP in (i) the agonist action of opiates and (ii) the expression of the abstinence syndrome, it follows that opiate dependence is a state of heightened potential activity of a neuronal cyclic AMP mechanism, initiated and maintained by the blockade of an adenylate cyclase. Various possible mechanisms are discussed by which this potential is heightened. New evidence is given that morphine and naloxone stimulate prostaglandin biosynthesis without mutual antagonism. Preliminary evidence also is given that (i) the formation of cyclic AMP is enhanced in brain homogenates from heroin-dependent rats, and (ii) an acidified ethylacetate extract of brains of morphine-dependent rats induces quasi-abstinence effects when injected into a lateral cerebral ventricle of naive rats.     

Prostaglandins, cyclic AMP and the biochemical mechanism of opiate agonist action.

The paper re-examines, in the light of recent evidence, the original proposition that the ability of opiates to inhibit the stimulation by E prostaglandins of cyclic AMP formation in rat brain homogenate represents a biochemical mechanism that could account for the analgesic and allied effects of these drugs. It is concluded that subsequent evidence largely supports the original proposition; but that this can now be made more definite. It is proposed that inhibition of an adenylate cyclase of morphine-sensitive neurones is the decisive biochemical consequence of the binding of an opiate agonist with its specific receptor.