Evidence that acetyl phosphate functions as a global signal during biofilm development

We used DNA macroarray analysis to identify genes that respond to the status of the intracellular acetyl phosphate (acP) pool. Genes whose expression correlated negatively with the ability to synthesize acP (i.e. negatively regulated genes) function primarily in flagella biosynthesis, a result consistent with observations that we published previously (Prüss and Wolfe, 1994, Mol Microbiol 12: 973-984). In contrast, genes whose expression correlated positively with the ability to synthesize acP (i.e. positively regulated genes) include those for type 1 pilus assembly, colanic acid (capsule) biosynthesis and certain stress effectors. To our knowledge, this constitutes the first report that these genes may respond to the status of the intracellular acP pool. Previously, other researchers have implicated flagella, type 1 pili, capsule and diverse stress effectors in the formation of biofilms. We therefore tested whether cells altered in their ability to metabolize acP could construct normal biofilms, and found that they could not. Cells defective for the production of acP and cells defective for the degradation of acP could both form biofilms, but these biofilms exhibited characteristics substantially different from each other and from biofilms formed by their wild-type parent. We confirmed the role of individual cell surface structures, the expression of which appears to correlate with acP levels, in fim or fli mutants that cannot assemble type 1 pili or flagella respectively. Thus, the information gained by expression profiling of cells with altered acP metabolism indicates that acP may help to co-ordinate the expression of surface structures and cellular processes involved in the initial stages of wild-type biofilm development.     

Model for food chain transfer and dose assessment in areas of the Slovak Republic

The radio-ecological model ECOSYS-87 developed for German ecological conditions, has been adapted to the conditions of two sub-regions in the Slovak Republic. In particular, the selection of plant varieties used for animal feeding, the plants’ growing cycles and harvests, the animal feeding practices and the human consumption rates were subjected to adaptation. Measurements of caesium and iodine radioactivity in soil, plants and animal products that have mainly been performed in the vicinity of the Bohunice nuclear power plant after the Chernobyl accident, are compared with the results of the adapted model. Data from various locations with dry, mixed or predominantly wet deposition show in general good agreement. Received: 10 April 2000 / Accepted: 23 October 2000     

Initiation of DNA replication by DNA polymerases from primers forming a triple helix

Despite extensive studies on oligonucleotide-forming triple helices, which were discovered in 1957, their possible relevance in the initiation of DNA replication remains unknown. Using sequences forming triple helices, we have developed a DNA polymerisation assay by using hairpin DNA templates with a 3′ dideoxynucleotide end and an unpaired 5′-end extension to be replicated. The T7 DNA polymerase successfully elongated nucleotides to the expected size of the template from the primers forming triple helices composed of 9–14 deoxyguanosine-rich residues. The triple helix-forming primer required for this reaction has to be oriented parallel to the homologous sequence of the hairpin DNA template. Substitution of the deoxyguanosine residues by N7 deazadeoxyguanosines in the hairpin of the template prevented primer elongation, suggesting that the formation of a triple helix is a prerequisite for primer elongation. Furthermore, DNA sequencing could be achieved with the hairpin template through partial elongation of the third DNA strand forming primer. The T4 DNA polymerase and the Klenow fragment of DNA polymerase I provided similar DNA elongation to the T7 polymerase–thioredoxin complex. On the basis of published crystallographic data, we show that the third DNA strand primer fits within the catalytic centre of the T7 DNA polymerase, thus underlying this new property of several DNA polymerases which may be relevant to genome rearrangements and to the evolution of the genetic apparatus, namely the DNA structure and replication processes.     

Crucial role of TNF-alpha in CD8 T cell-mediated elimination of 3LL-A9 Lewis lung carcinoma cells in vivo

The role of perforin, IFN-gamma, and TNF-alpha in anti-tumor CD8 T cell immunity was examined in a new tumor model using a CD8 T cell epitope (GP33) derived from lymphocytic choriomeningitis virus as a tumor-associated Ag. In contrast with parental 3LL-A9 (A9) Lewis lung carcinoma cells that progressively grow in C57BL/6 mice, s.c. injection of GP33-transfected A9GP33 tumor cells induced a protective GP33-specific CD8 T cell response that led to complete tumor cell elimination. Tumor regression was dependent on perforin, IFN-gamma, or TNF-alpha, because A9GP33 tumors developed in mice deficient in one of these genes. A9GP33 tumors arising in perforin- and IFN-gamma-deficient mice represented GP33 Ag-loss variants, demonstrating that GP33-specific CD8 T cells from these mice were able to exert an Ag selection pressure. In contrast, tumor cells growing in TNF-alpha knock-out mice still expressed the tumor-associated GP33 peptide despite the presence of activated GP33-specific CD8 T cells. These findings provide evidence for a crucial role of TNF-alpha in A9 tumor cell elimination by CD8 T cells in vivo.     

Ciboulot regulates actin assembly during Drosophila brain metamorphosis

A dynamic actin cytoskeleton is essential for the remodeling of cell shape during development, but the specific roles of many actin partners remain unclear. Here we characterize a novel actin binding protein, Ciboulot (Cib), which plays a major role in axonal growth during Drosophila brain metamorphosis. Loss of Cib function leads to axonal growth defects in the central brain, while overexpression of the gene during development leads to overgrown projections. The Cib protein displays strong sequence similarity to beta-thymosins but has biochemical properties like profilin: the Cib-actin complex participates in actin filament assembly exclusively at the barbed end, and Cib enhances actin-based motility in vitro. Genetic experiments show that Cib and the Drosophila profilin protein Chickadee (Chic) cooperate in central brain metamorphosis.     

Characterization of processes responsible for the distinct effect of herbicides DCMU and BNT on Photosystem II photoinactivation in cells of the cyanobacterium Synechococcus sp. PCC 7942

Light-induced modification of Photosystem II (PS II) complex was characterized in the cyanobacterium Synechococcus sp. PCC 7942 treated with either DCMU (a phenylurea PS II inhibitor) or BNT (a phenolic PS II inhibitor). The irradiance response of photoinactivation of PS II oxygen evolution indicated a BNT-specific photoinhibition that saturated at relatively low intensity of light. This BNT-specific process was slowed down under anaerobiosis, was accompanied by the oxygen-dependent formation of a 39 kDa D1 protein adduct, and was not related to stable Q_A reduction or the ADRY effect. In the BNT-treated cells, the light-induced, oxygen-independent initial drop of PS II electron flow was not affected by formate, an anion modifying properties of the PS II non-heme iron. For DCMU-treated cells, anaerobiosis did not significantly affect PS II photoinactivation, the D1 adduct was not observed and addition of formate induced similar initial decrease of PS II electron flow as in the BNT-treated cells. Our results indicate that reactive oxygen species (most likely singlet oxygen) and modification of the PS II acceptor side are responsible for the fast BNT-induced photoinactivation of PS II. This revised version was published online in June 2006 with corrections to the Cover Date.     

Probiotic strain <Emphasis Type="Italic">Bacillus subtilis</Emphasis> CU1 stimulates immune system of elderly during common infectious disease period: a randomized,double-blind placebo-controlled study

Background

Bacillus probiotics health benefits have been until now quite poorly studied in the elderly population. This study aimed to assess the effects of Bacillus subtilis CU1 consumption on immune stimulation and resistance to common infectious disease (CID) episodes in healthy free-living seniors.

Results

One hundred subjects aged 60–74 were included in this randomized, double-blind, placebo-controlled, parallel-arms study. Subjects consumed either the placebo or the probiotic (2.10⁹ B. subtilis CU1 spores daily) by short periodical courses of 10 days intermittently, alternating 18-day course of break. This scheme was repeated 4 times during the study. Symptoms of gastrointestinal and upper/lower respiratory tract infections were recorded daily by the subjects throughout the study (4 months). Blood, saliva and stool samples were collected in a predefined subset of the first forty-four subjects enrolled in the study. B. subtilis CU1 supplementation did not statistically significantly decrease the mean number of days of reported CID symptoms over the 4-month of study (probiotic group: 5.1 (7.0) d, placebo group: 6.6 (7.3) d, P?=?0.2015). However, in the subset of forty-four randomized subjects providing biological samples, we showed that consumption of B. subtilis CU1 significantly increased fecal and salivary secretory IgA concentrations compared to the placebo. A post-hoc analysis on this subset showed a decreased frequency of respiratory infections in the probiotc group compared to the placebo group.

Conclusion

Taken together, our study provides evidence that B. subtilis CU1 supplementation during the winter period may be a safe effective way to stimulate immune responses in elderly subjects.