Intranasal vaccination with a recombinant vesicular stomatitis virus expressing cottontail rabbit papillomavirus L1 protein provides complete protection against papillomavirus-induced disease

Immunizations with live recombinant vesicular stomatitis viruses (rVSV) expressing foreign viral proteins have successfully protected animals from challenges with several heterologous viruses. We developed an rVSV expressing the major capsid protein (L1) of cottontail rabbit papillomavirus (CRPV) and tested the efficacy of protection following CRPV challenge. An rVSV expressing L1 of CRPV (VSV-L1) was characterized for the protective ability afforded by intranasal, intradermal, or intramuscular vaccination in rabbits subsequently challenged with CRPV. Protein expression of L1 in VSV-L1 was confirmed by radioimmunoprecipitation assays. Nuclear localization of L1 was demonstrated by indirect immunofluorescence assays. Immunized rabbits elicited significant VSV neutralization and VLP-L1 enzyme-linked immunosorbent assay titers. VSV-L1 vaccination was not associated with weight loss or any other adverse clinical signs in the rabbit model. VSV shedding in nasal secretions occurred in some rabbits, peaking at 4 to 6 days after intranasal vaccination, with no further shedding after day 6. Specific humoral immunity to the L1 protein was consistently seen after a single VSV-L1 vaccination when administered through an intradermal or intramuscular route or after a boost via the intranasal route. Rabbits were completely protected from CRPV-induced papillomas after VSV-L1 vaccination and boost given intranasally or intramuscularly. Vaccination with VSV-L1 is a novel approach to prevent papillomavirus-induced disease and demonstrates a potential strategy for developing a human papillomavirus vaccine that can be given without injection.     

Mutation of rpiA in Enterobacter cloacae decreases seed and root colonization and biocontrol of damping-off caused by Pythium ultimum on cucumber

Strains of Enterobacter cloacae show promise as biocontrol agents for Pythium ultimum-induced damping-off on cucumber and other crops. E. cloacae A145 is a mini-Tn5 Km transposon mutant of strain 501R3 that was significantly reduced in suppression of damping-off on cucumber caused by P. ultimum. Strain A145 was deficient in colonization of cucumber, sunflower, and wheat seeds and significantly reduced in colonization of corn and cowpea seeds relative to strain 501R3. Populations of strain A145 were also significantly lower than those of strain 501R3 at all sampling times in cucumber, wheat, and sunflower rhizosphere. Populations of strain A145 were not detectable in any rhizosphere after 42 days, while populations of strain 501R3 remained at substantial levels throughout all experiments. Molecular characterization of strain A145 indicated mini-Tn5 Km was inserted in a region of the E. cloacae genome with a high degree of DNA and amino acid sequence similarity to rpiA, which encodes ribose-5-phosphate isomerase. In Escherichia coli, RpiA catalyzes the interconversion of ribose-5-phosphate and ribulose-5-phosphate and is a key enzyme in the pentose phosphate pathway. Ribose-5-phosphate isomerase activity in cell lysates from strain A145 was approximately 3.5% of that from strain 501R3. In addition, strain A145 was a ribose auxotroph, as expected for an rpiA mutant. Introduction of a 1.0-kb DNA fragment containing only the rpiA homologue into strain A145 restored ribose phosphate isomerase activity, prototrophy, seedling colonization, and disease suppression to levels similar to those associated with strain 501R3. Experiments reported here indicate a key role for rpiA and possibly the pentose phosphate pathway in suppression of damping-off and colonization of subterranean portions of plants by E. cloacae.     

Diabetic neuropathy and surface sway-referencing disrupt somatosensory information for postural stability in stance

In order to determine the type of somatosensory information for postural control that is most affected by neuropathy, we compared the relative effects of three methods of sway-referencing the surface in a group of subjects with profound loss of somatosensory function associated with sensory polyneuropathy from diabetes with age-matched control subjects. Sway-referencing disrupted somatosensory feedback for postural control by servo-controlling the dorsi- and plantar-flexion rotation of the support surface in proportion to anterior-posterior excursion of (1) ankle angle, (2) center of body mass (CoM) angle or (3) filtered center of pressure (CoP). Postural sway in subjects with somatosensory loss was significantly larger than normal on a firm surface but not on the sway-referenced surfaces, suggesting that sway-referencing disrupts somatosensory information for postural control already disrupted by neuropathy. Control subjects standing on any sway-referenced surface swayed significantly more than neuropathy subjects who stood on a firm surface, suggesting that sway-referencing disrupts more somatosensory information than disrupted by severe neuropathy. CoP sway-referencing was less sensitive than ankle or CoM sway-referencing for distinguishing postural sway in subjects with somatosensory loss from age-matched control subjects. Given that filtered CoP sway-referencing disrupts the ability to utilize somatosensory information related to surface reactive force to a greater extent than the other two methods of sway-referencing, then these results support the hypothesis that subjects with diabetic peripheral neuropathy have lost more CoP information, than ankle or CoM angle information, for controlling postural sway in stance.     

An NMR investigation of the conformational equilibria of 2-(2'-pyridyl)ethylphosphonic acid in several solvents

Vicinal proton-proton NMR couplings have been used to investigate whether the position of conformational equilibria is determined by intramolecular N-H hydrogen bonding for 2-(2'-pyridyl)ethylphosphonic acid 1 in its various possible ionization states in water, methanol, ethanol, and dimethyl sulfoxide (DMSO). With 1 in the form of its monoanion and dianion, the trans is favored, with the dianion being more trans than the monoanion for a given solvent, probably as the result of steric effects, possibly enhanced by repulsive electrostatic effects between the negatively charged phosphonic group and the lone pair on the pyridine nitrogen. For 1 and its conjugate acid, the gauche amounts, respectively, to 43% and 45% in water, 66% and 51% in methanol, 66% and 64% in ethanol, and 29% and 49% in DMSO. For these latter two species, electrostatic, steric, and hydrogen bonding-effects are all likely to play a role in determining the conformational equilibria.     

A germline-specific gap junction protein required for survival of differentiating early germ cells

Germ cells require intimate associations and signals from the surrounding somatic cells throughout gametogenesis. The zero population growth (zpg) locus of Drosophila encodes a germline-specific gap junction protein, Innexin 4, that is required for survival of differentiating early germ cells during gametogenesis in both sexes. Animals with a null mutation in zpg are viable but sterile and have tiny gonads. Adult zpg-null gonads contain small numbers of early germ cells, resembling stem cells or early spermatogonia or oogonia, but lack later stages of germ cell differentiation. In the male, Zpg protein localizes to the surface of spermatogonia, primarily on the sides adjacent to the somatic cyst cells. In the female, Zpg protein localizes to germ cell surfaces, both those adjacent to surrounding somatic cells and those adjacent to other germ cells. We propose that Zpg-containing gap junctional hemichannels in the germ cell plasma membrane may connect with hemichannels made of other innexin isoforms on adjacent somatic cells. Gap junctional intercellular communication via these channels may mediate passage of crucial small molecules or signals between germline and somatic support cells required for survival and differentiation of early germ cells in both sexes.     

A role for nuclear localised proteasomes in mediating auxin action

A number of important cellular events in animals and yeast are regulated by protein degradation, and it is becoming apparent that such regulated proteolysis is involved in many facets of plant physiology and development. We have investigated the role of protein degradation by proteasomes in plants using NtPSA1, a tobacco gene that is predominantly expressed in young developing tobacco tissues and has extensive homology to yeast and human alpha-type proteasome subunit genes. The NtPSA1 cDNA was used to complement a lethal mutation of the yeast PRC1 alpha subunit gene indicating that NtPSA1 encodes a functional proteasome subunit, and transient expression of an NtPSA1::GUS protein fusion in onion cells confirmed that the nuclear localisation signal that is present in the NtPSA1 peptide sequence is active in plant cells. Plants transformed with an antisense NtPSA1 gene had reduced levels of NtPSA1 mRNA and exhibited reduced apical dominance. In addition, these low NtPSA1 plants displayed several morphological defects associated with auxin resistance such as reduced stamen length, and showed increased tolerance to high concentrations of auxin. These results support a role for nuclear localised proteasomes in floral development and auxin responses.     

Synthesis and structure-activity relationships of M(2)-selective muscarinic receptor ligands in the 1-[4-(4-arylsulfonyl)-phenylmethyl]-4-(4-piperidinyl)-piperazine family

The synthesis and muscarinic binding properties of compounds based on the 1-[4-(4-arylsulfonyl)phenylmethyl]-4-(1-aroyl-4-piperidinyl)-piperazine skeleton are described. For compounds, substituted with appropriately configured methyl groups at the benzylic center and at the piperazine 2-position, high levels of selective, M(2) subtype affinity could be obtained, particularly when the terminal N-aroyl residue was ortho-substituted.     

The pH of the Plasmodium falciparum digestive vacuole: holy grail or dead-end trail?

The maintenance of acidic pH in the digestive vacuole of the malaria parasite is thought to be crucial to the digestion of host cell haemoglobin and the subsequent process of heme detoxification. It may also be important in the mode of action of chloroquine and in the mechanism of resistance to the drug. Obtaining a definitive measurement of digestive vacuole pH has been surprisingly difficult. Some of the techniques for the measurement of pH in acid vesicles are outlined here along with some key aspects that are specific to malaria parasites. The use of acridine orange and dextran-tagged dyes as probes for the measurement of digestive vacuole pH has proved problematic, yet some surprising findings have emerged from work with these compounds.