Expression of Shigella sonnei lipopolysaccharide in Vibrio cholerae

Making use of a newly designed mobilizable suicide vector, the genetic determinants encoding Shigella sonnei lipopolysaccharide (LPS) were stably integrated into the chromosome of the live attenuated Vibrio cholerae vaccine strain CVD103-HgR. Expression studies showed that the production of complete S. sonnei O-polysaccharide (O-PS)-bearing LPS was limited in bivalent recombinant strains that were also proficient in the synthesis of the host-encoded Inaba O-PS. Conversely, high amounts of LPS carrying S. sonnei O-PS are produced in monovalent Inaba-deficient derivatives, even in those strains which do not co-express the compatible R1 LPS core. Thus, the non-enterobacterial V. cholerae LPS core efficiently acts as a receptor for covalent binding of S. sonnei O-PS provided that competition with the host O-PS is avoided. Expression of the R1 core interferes with cell division in recombinant V. cholerae without affecting other physiological properties of vaccine strain CVD103-HgR. Both monovalent and bivalent strains stimulated high serum-antibody titres specific for their respective O-serotype(s) when administered to rabbits. The potential of V. cholerae as an expression carrier for heterologous O-serotypes is discussed.     

Temperature-pulse-induced "memory" in Bacillus subtilis macrofibers and a role for protein(s) in the left-handed-twist state

Macrofibers in steady-state growth at one temperature were subjected to pulses of various durations at a temperature at which the opposite helix hand would form and then returned to the initial temperature. In an upshift pulse (20 to 48 degrees C), at least 3 min of incubation was required to induce a transient inversion that occurred later after return to 20 degrees C. Longer pulses resulted in shorter delays in onset of the transient inversion. This "memory" of a brief high-temperature pulse suggests that even a small amount of material can influence the twist of the entire macrofiber. Similar results were found for temperature downshift pulses corresponding to the opposite inversion. Adding chloramphenicol during the temperature pulse blocked the establishment of memory associated with the right-to-left inversion but not that associated with left-to-right inversion. In contrast, inhibiting peptidoglycan synthesis with D-cycloserine during the temperature pulse did not prevent establishment of memory. Inhibiting protein synthesis in mutants fixed as left-handed structures over the entire temperature range induced conversion to right-handedness but did not affect mutants fixed as right-handed structures. Adding protease to either live or formaldehyde-killed macrofibers always induced rotations of right-handed orientation. Steady-state growth in the presence of protease was found to shift the initial macrofiber twist towards the right-hand end of the twist spectrum. The phenomenon was observed in several mutants with different initial twists.     

The T4 mot protein functions as part of a pre-replicative DNA-protein complex

Middle-mode RNA synthesis in T4-infected cells takes place before replication of phage DNA commences. What distinguishes it from early-mode RNA synthesis is that initiation of middle RNA depends on T4-coded proteins, in particular on the mot gene product. mot protein is localized in a DNA-protein complex which forms during the first few minutes of infection. All of the cell's mot protein is bound in this complex, and it continues to be bound long after the synthesis of mot protein has stopped. When we infect Escherichia coli with T4 carrying a temperature-sensitive mutation in the mot gene, we find a correlation between the physiology of this mot mutant and the amount of mot protein bound in the DNA-protein complex. Although there is some host RNA polymerase in the complex, mot protein does not seem to bind to this enzyme. Two other T4-coded proteins, of molecular weights 17,600 and 15,000, are also found in the pre-replicative DNA-protein complex. One of these, p17,600, is coded for by a 750-base pair region located between genes 39 and 56; p17,600 appears to be the recently described motB gene product. The other protein, p15,000, is not an RNA polymerase-binding protein; it is characterized by its strong binding to the DNA-protein complex.     

Operant conditioning of scratching in lemurs

An adult femaleLemur catta and an adult femaleEulemur fulvus were given edible rewards for scratching. Both subjects learned to scratch in order to obtain the rewards, showed diminished rates of scratching during periods of extinction, and learned to scratch preferentially with one foot when required. TheLemur catta subject was more responsive to the changing experimental conditions than theEulemur fulvus. The conditionability of scratching in primates does not appear to be directly related to the widespread occurrence of scratching in simian social contexts.     

Phylogeny of Greya (Lepidoptera: Prodoxidae), based on nucleotide sequence variation in mitochondrial cytochrome oxidase I and II: congruence with morphological data

The phylogeny of Greya Busck (Lepidoptera: Prodoxidae) was inferred from nucleotide sequence variation across a 765-bp region in the cytochrome oxidase I and II genes of the mitochondrial genome. Most parsimonious relationships of 25 haplotypes from 16 Greya species and two outgroup genera (Tetragma and Prodoxus) showed substantial congruence with the species relationships indicated by morphological variation. Differences between mitochondrial and morphological trees were found primarily in the positions of two species, G. variabilis and G. pectinifera, and in the branching order of the three major species groups in the genus. Conflicts between the data sets were examined by comparing levels of homoplasy in characters supporting alternative hypotheses. The phylogeny of Greya species suggests that host-plant association at the family level and larval feeding mode are conservative characters. Transition/transversion ratios estimated by reconstruction of nucleotide substitutions on the phylogeny had a range of 2.0-9.3, when different subsets of the phylogeny were used. The decline of this ratio with the increase in maximum sequence divergence among taxa indicates that transitions are masked by transversions along deeper internodes or long branches of the phylogeny. Among transitions, substitutions of A-->G and T-->C outnumbered their reciprocal substitutions by 2-6 times, presumably because of the approximately 4:1 (77%) A+T-bias in nucleotide base composition. Of all transversions, 73%-80% were A<-->T substitutions, 85% of which occurred at third positions of codons; these estimates did not decrease with an increase in maximum sequence divergence of taxa included in the analysis. The high frequency of A<-->T substitutions is either a reflection or an explanation of the 92% A+T bias at third codon positions.