Ethylene and in vitro rooting of hazelnut (Corylus avellana) cotyledons

Ethylene may be one of the many factors that play a role in rooting. However, in some studies ethylene promoted rooting, while in others it was inhibitory or had no effect. Using cotyledons of hazelnut ( Corylus avellana L. cv. Casina) observations were made of the effect of ethylene precursors on adventitious root formation. l-methionine (Met) or 1-aminocyclopropane-1-carboxylic acid (ACC) added to a standard indole-3-butyric acid (IBA)-kinetin-containing medium did not enhance rooting, while 2-chloroethylphosphonic acid (CEPA) did. The ethylene inhibitor, aminoethoxyvinylglycine (AVG), inhibited root formation, but its effect was reversed by ACC when cotyledonary segments were transferred to rhizogenic medium plus ACC at day 10. Ethylene production by cotyledons cultured on rhizogenic medium or rhizogenic medium plus CEPA was high at the beginning of rooting. Thus, the wound-induced ethylene is a key stimulatory factor in the formation of root primordia. The data support the hypothesis that ethylene plays a positive role in root formation.     

Four cases of direct ion channel gating by cyclic nucleotides

Four different nucleotide-gated ion channels are discussed in terms of their biophysical properties and their importance in cell physiology. Channels activated directly by cGMP are present in vertebrate and invertebrate photoreceptors. In both cases cGMP increases the fraction of time the channel remains in the open state. At least three cGMP molecules are involved in channel opening in vertebrate photoreceptors and the concentration of the cyclic nucleotide to obtain the half maximal effect is about 15 µM. The light-dependent channel of both vertebrates and invertebrates is poorly cation selective. The vertebrate channel allows divalent cations to pass through 10–15-fold more easily than monovalent ions. In agreement with their preference for divalent cations, this channel is blocked byl-cis Dialtazem, a molecule that blocks certain types of calcium channels. In olfactory neurons a channel activated by both cAMP and cGMP is found and, as in the light-dependent channel, several molecules of the nucleotide are needed to open the channel with a half maximal effect obtained in the range of 1–40 µM. The channel is poorly cationic selective. A K⁺ channel directly and specifically activated by cAMP is found inDrosophila larval muscle. At least three cAMP molecules are involved in the opening reaction. Half-maximal effect is obtained at about 50 µM. This channel is blocked by micromolar amount of tetraethylammonium applied internally. Interestingly, this channel has a probability of opening 10–20-fold larger in the mutantdunce, a mutant that possesses abnormally elevated intracellular cAMP level, than in the wild type.     

Cell type-specific gene expression in the Drosophila melanogaster male accessory gland.

The accessory gland of the male Drosophila melanogaster plays a vital role in reproduction. This secretory organ synthesizes products that are transferred to the female and are necessary to elicit the proper physiological and behavioral responses in the female. The accessory gland is composed of two morphologically distinct secretory cell types, the main cells and the secondary cells. Previous studies identified some genes expressed in main cells or in all accessory gland cells. In this paper we use P-element mediated enhancer traps to examine gene expression in the accessory gland. We show that, in addition to genes expressed in main cells only or in all accessory gland secretory cells, there are genes expressed specifically in secondary cells. Each cell type is uniform in the expression of its genes. Our results demonstrate that the two cell types are not only morphologically distinct but also biochemically distinct. We also show that the two cell types differ in their regulation of gene expression in response to mating activity.     

Molecular and symbiotic characterization of exopolysaccharide-deficient mutants of Rhizobium tropici strain CIAT899

We studied the symbiotic behaviour of 20 independent Tn5 mutants of Rhizobium tropici strain CIAT899 that were deficient in exopolysaccharide (EPS) production. The mutants produced non-mucoid colonies, were motile, grew in broth cultures at rates similar to those of the parent, and produced significantly less EPS than did CIAT899 in broth culture. A genomic library of strain CIAT899, constructed in pLA2917, was mobilized into all of the mutants, and cosmids that restored EPS production were identified. EcoRI restriction digests of the cosmids revealed nine unique inserts. Mutant complementation and hybridization analysis showed that the mutations affecting EPS production fell into six functional and physical linkage groups. On bean, the mutants were as efficient in nodulation and as effective in acetylene reduction as strain CIAT899, induced a severe interveinal chlorosis, and all but one were less competitive than CIAT899. On siratro, CIAT899 induced nodules that were ineffective in acetylene reduction, whereas the EPS-deficient mutants induced effective nodules. Microscopic examination of thin sections showed that nodules from both siratro and bean plants inoculated with either CIAT899 or an EPS-deficient mutant contained infected cells. These data indicate that EPS is not required for normal nodulation of bean by R. tropici, that it may contribute to competitiveness of R. tropici on bean, and that the loss of EPS production is accompanied by acquisition of the ability to reduce acetylene on siratro.     

Effects of phosphorothioate capping on antisense oligonucleotide stability, hybridization and antiviral efficacy versus herpes simplex virus infection.

Efforts have been made to improve the biological stability of phosphodiester (PO) oligonucleotides by the addition of various modifications to either the 3', 5' or both the 3' and 5' ends of an oligonucleotide. ISIS 1080, a phosphorothioate (PS) 21-mer oligonucleotide complementary to the internal AUG codon of UL13 mRNA in HSV-1, reduces the infectious yield of HSV-1 in HeLa cells to 9.0% +/- 11%. PO analogs of ISIS 1080 containing three PS linkages placed on the 3' (ISIS 1365), 5' (ISIS 1370), both the 3' and 5' (ISIS 1364) ends or with four linkages in the middle (ISIS 1400) demonstrated reduced antiviral efficacy compared to fully PS ISIS 1080. Thermal denaturation profiles demonstrated that these oligonucleotides hybridized to complementary DNA or RNA with equivalent binding affinities. All were able to support E. coli RNAse H cleavage of the HSV mRNA to which they were targeted. The stability of the congeners in cell culture medium containing 10% fetal calf serum (FCS), HeLa cytosolic extract, HeLa nuclear extract and in intact HeLa cells revealed that ISIS 1080 was most resistant to nucleolytic digestion through 48 hours. Partial PS oligonucleotides exhibited increased degradation compared to the fully thioated oligonucleotide by exonuclease activity in FCS and endonuclease activity in cell extracts or intact cells. Thus, the reduced efficacy of partial compared to fully PS oligonucleotides against HSV-1 in HeLa cells may result from increased degradation of the mixed PO/PS oligonucleotides.     

Investigation of cis/trans proline isomerism in a multiply occurring peptide fragment from human salivary proline-rich glycoprotein.

The solution-state conformations of eight proline-containing peptide fragments found in human salivary proline-rich glycoprotein (PRG) were investigated in 2 x distilled water (treated with metal ion chelating resin) using 13C-nuclear magnetic resonance (NMR) and circular dichroism (CD) spectroscopy. The peptide sequences and acronyms were as follows: PRG9-2 = NH2-G(1)-P(2)-CONH2, PRG9-3 = NH2-G(1)P(2)-P(3)-CONH2, PRG9-4 = NH2-G(1)-P(2)-P(3)-P(4)-CONH2, PRG9-5 = NH2-G(1)-P(2)-P(3)-P(4)-H(5)-CONH2, PRG9-6 = NH2-G(1)-P(2)-P(3)-P(4)-H(5)-P(6)-CONH2, PRG9-7 = NH2-G(1)-P(2)-P(3)-P(4)-H(5)-P(6)-G(7)-CONH2, PRG9-8 = NH2-G(1)-P(2)-P(3)-P(4)-H(5)-P(6)-G(7)-K(8)-CONH2 and PRG9-9 = NH2-G(1)-P(2)-P(3)-P(4)-H(5)-P(6)-G(7)-K(8)-P(9)-CONH2. Sequence-specific resonance assignments from the 13C-NMR spectra indicated that the trans proline isomer dominated the conformations of the peptides. CD results clearly showed the presence of the poly-L-proline II helix as the major conformation in PRG9-3----PRG9-5, supplemented by beta- and/or gamma-turns in PRG9-6----PRG9-9. These data suggest that in "metal free" water, native PRG could contain several small poly-L-proline II helices along with beta- and/or gamma-turns. Since proline is the major amino acid present in native PRG, these localized conformations may contribute to PRG's global conformation and act as a primary force in determining its biological activities.     

Oxidation of Ferrous Iron and Elemental Sulfur by Thiobacillus ferrooxidans

The oxidation of ferrous iron and elemental sulfur by Thiobacillus ferrooxidans that was absorbed and unabsorbed onto the surface of sulfur prills was studied. Unadsorbed sulfur-grown cells oxidized ferrous iron at a rate that was 3 to 7 times slower than that of ferrous iron-grown cells, but sulfur-grown cells were able to reach the oxidation rate of the ferrous iron-adapted cells after only 1.5 generations in a medium containing ferrous iron. Bacteria that were adsorbed to sulfur prills oxidized ferrous iron at a rate similar to that of unadsorbed sulfur-grown bacteria. They also showed the enhancement of ferrous iron oxidation activity in the presence of ferrous iron, even though sulfur continued to be available to the bacteria in this case. An increase in the level of rusticyanin together with the enhancement of the ferrous iron oxidation rate were observed in both sulfur-adsorbed and unadsorbed cells. On the other hand, sulfur oxidation by the adsorbed bacteria was not affected by the presence of ferrous iron in the medium. When bacteria that were adsorbed to sulfur prills were grown at a higher pH (ca. 2.5) in the presence of ferrous iron, they rapidly lost both ferrous iron and sulfur oxidation capacities and became inactive, apparently because of the deposition of a jarosite-like precipitate onto the surface to which they were attached.     

Phenolsulphonphthalein conjugation and biliary excretion in the rat: influence of phenobarbital and 3-methylcholanthrene

The effect of phenobarbital and 3-methylcholanthrene pretreatment on the biliary excretion of phenolsulphonphthalein (PSP) was investigated in male Wistar rats. The dye was injected at a single dose of 200 mumol/kg body wt. About 20% of the compound was excreted as a glucuronide in the controls, the liver UDP-glucuronyltransferase activity toward PSP being 0.064 +/- 0.005 nmol.min-1.mg protein-1. Treatment for two weeks with phenobarbital (354 mumol.kg body wt-1.day-1) caused a transient increase in conjugated and unconjugated PSP excretion, but glucuronyltransferase activity was not modified. 3-Methylcholanthrene pretreatment for 4 days (75 mumol.kg body wt-1.day-1) also enhanced biliary excretion of the dye, but the increase corresponded only to the glucuronide and glucuronyltransferase activity was significantly enhanced by 20%. Our data indicate that not only the rate of biotransformation but also other factors could be responsible for increased PSP biliary excretion following administration of microsomal enzyme inducers.