Differential sensitivity to tributyltin of cytochrome-containing and cytochrome-deficient cells of Escherichia coli SASX76

The effect of tributyltin (TBT) chloride on the growth of cytochrome-deficient and cytochrome-containing cells of Escherichia coli SASX76 was examined. The former cells were found to be at least 20 times more sensitive to TBT. It is proposed that the differential sensitivity of these two cell types to the biocide, TBT, may be due to a different mode of energy generation by cytochrome-deficient and cytochrome-sufficient cells. In addition to the energy state, the pH change caused by the presence and absence of cytochromes which occurred during growth also resulted in a differential sensitivity of these cells.     

Oxidative stress in the moderately halophilic bacteriumDeleya halophila: Effect of NaCl concentration

The sensitivity ofDeleya halophila to oxidative stress caused by hydrogen peroxide (H₂O₂) was found to vary, depending on the NaCl concentration of the growth medium. Pretreatment of the bacteria at a low concentration of H₂O₂ (50 M) protected the cells against the lethal effects of higher levels (1–2 mM) of H₂O₂. Exposure ofD. halophila cells to 50 M H₂O₂ resulted in the induction of several proteins (hydrogen peroxide-inducible proteins, hips). However, the kinetics of induction, the extent of induction and the number of hips appear to be influenced by the salt concentration of the growth medium. Five of the hips exhibited apparent molecular masses identical to those of five heat shock proteins (hsps).     

Hydroxyproline-rich glycoproteins in plant reproductive tissues: structure, functions and regulation

The plant reproductive process of pollination involves a series of interactions between the male gametophyte (the pollen grain or pollen tube) and extracellular matrix (ECM) molecules secreted by different cell types along the pollen tube growth pathway in the female organ, the pistil. These interactions are believed to signal and regulate the pollen tube growth process to effect successful delivery of the sperm cells to the ovules where fertilization takes place. Hydroxyproline-rich glycoproteins secreted by plant cells are believed to play a broad range of functions, ranging from providing structural integrity to mediating cell-cell interactions and communication. The pistil and pollen tube ECM is enriched in these highly glycosylated proteins. Our discussions here will focus on a number of these proteins for which most information has been available, from Nicotiana tabacum, its self-incompatible relative N. alata, and Zea mays. In addition, the regulation of the synthesis and glyco-modification of one of these proteins, TTS (transmitting tissue-specific) protein from N. tabacum will be discussed in the light of how differential glycosylation may be used to regulate molecular interactions within the ECM.     

Somatic variation in micropropagated clones ofCichorium intybus

Summary An experiment aiming to establish whether temporary or persistet somatic variation can arise in clones obtained in vitro by micropropagation has been performed onCichorium intybus. The results point out that persistent modifications of the phenotype can be observed after six cycles of cloning and that when the length of the cloning period is varied these appear to be differential responses of the same genotype to the micropropagation.     

DNG1, a Dictyostelium homologue of tumor suppressor ING1 regulates differentiation of Dictyostelium cells

dng1 is a Dictyostelium homologue of the mammalian tumor suppressor ING gene. DNG1 protein localizes in the nucleus, and has a highly conserved PHD finger domain found in chromatin-remodeling proteins. Both dng1 disruption and overexpression impaired cell proliferation. In dng1-null cells, the progression of differentiation was delayed in a cell-density-dependent manner, and many tiny aggregates were formed. Exogenously applied cAMP pulses reversed the inhibitory effect caused by dng1 disruption on the aggregation during early development, but formation of tiny aggregates was not restored. dng1-overexpressing cells acquired the ability to undergo chemotaxis to cAMP earlier and exhibited enhanced differentiation. These phenotypes were found to be coupled with altered expressions of early genes such as cAMP receptor 1 (car1) and contact site A (csA). Furthermore, disordered histone modifications were demonstrated in dng1-null cells. These results suggest a regulatory role of dng1 in the transition of cells from growth to differentiation.Received 29 December 2004; received after revision 24 May 2005; accepted 26 May 2005     

Osmotic adaptation of moderately halophilic methanogenic Archaeobacteria,and detection of cytosolicN,N-dimethylglycine

Methanohalophilus mahii SLP andMethanohalophilus halophilus Z-7982, two closely-related, moderately halophilic, methylotrophic methanogens, were tested for their adaptation to saline conditions. They grew in a wider range of salinities than previously reported, in a defined medium with as little as 0.1 M NaCl, and with a high as 4.0 M NaCl forM. halophilus and 4.5 M NaCl forM. mahii. Fastest growth occurred with 1.5 M NaCl forM. mahii and 1.0 M NaCl forM. halophilus. M. mahii also grew in media in which NaCl was replaced by sucrose or KCl as osmolytes up to the osmolal equivalent of 2 and 2.5 M NaCl (these media contained other sodium salts totaling about 0.1 M Na⁺). In media with either sucrose of KCl replacing NaCl,M. mahii grew fastest at osmolalities approximately equiosmolal to 1 M NaCl.M. mahii not only grew well at a wide range of osmosities, it also tolerated rapid shifts in osmolality. Cells subjected to a rapid 10-fold hypertonic shift resumed growth without a prolonged lag. When cells were subjected to a rapid 10-fold hypotonic shift, 90% of cells lysed, but the remaineder continued to swell with little further lysis during the next 45 min. Surviving cells resumed growth.Methanohalophilus strains grown in defined medium had low cytosolic Na⁺ concentrations; K⁺ concentrations were as high as 0.35 M. Organic osmotica in the cytosol include glycine betaine and larger amounts of N,N-dimethylglycine.     

Two-state model ofParamecium bursaria thigmotaxis

A theoretical framework has been developed for analysis of the interaction betweenParamecium bursaria and a glass surface. Adhesion to and detachment from a solid substrate were considered in the model as transitions between alternative states in cell behavior: (a) swimming, and (b) motionless (positive thigmotactic) state. According to the model and experimental data, a change in the fraction of swimming cells is described by a negative exponential course. The proposed model allows positive thigmotaxis, generally referred to in the literature simply as thigmotaxis, to be considered as the rate-constant of transition into the motionless state. This approach permits quantitative determination of thigmotaxis, and reveals its dependence on the phase of culture growth and the type of medium surrounding the cells. In the mineral maintenance solution, paramecia from the stationary phase of growth swim more slowly than those in the logarithmic phase of growth, and show enhanced thigmotaxis. However, a general relationship between thigmotaxis and swimming speed was not established.     

Measuring toxicity in marine environments: critical appraisal of three commonly used methods

Toxicity quantification is important in environmental monitoring, in the field of natural products, and in chemical ecology. The sensitivity and precision of three commonly used methods detecting toxicity in marine environments were compared, using the toxic marine spongeCrambe crambe as a test organism. The paper disk diffusion method (run with marine bacteria) showed the least sensitivity and did not permit toxicity levels to be quantified. The sea urchin and the MICROTOX® tests showed greater sensitivity, and the latter had the higher precision. The relative performance of these methods is discussed. It is concluded that the MICROTOX® bioassay displays the best characteristics for toxicity quantification.     

Phosphate transport inClaviceps sp. strain SD-58

Summary Phosphate uptake inClaviceps sp. strain SD-58 was found to be linear for 20 min, proportional to cell density in mg/ml, energy dependent, and taking place against a concentration gradient with a K_m value of 45.45×10^–5 M. Osmotic shock treatment to the cell caused a reduction in phosphate uptake associated with the release of binding protein. Partial restoration of uptake was observed on incubation of osmotically shocked cells with shock fluid. The results are discussed with reference to the effect of phosphate on alkaloid synthesis inClaviceps sp. strain SD-58.     

Shoot regeneration fromAgrobacterium tumefaciens-induced tumors of a tropical timber tree,Fagraea fragrans

Summary Agrobacterium tumefaciens A208 with nopaline plasmid pTiT37 was used to obtain stem tumors on plantlets ofFagraea fragrans grown in vitro. Bacterial elimination and tissue proliferation were simultaneously achieved by growing tumors on cefatoxime medium. After some tissue growth the shoots regenerated. An examination of these showed the presence of nopaline, indicating genetic transformation by T-DNA.     

Inhibition of respiratory oxidation of reduced sulfur compounds by intact cells ofThiobacillus denitrificans (strain RT) grown on thiosulfate

Thiobacillus denitrificans strain RT, an obligate sulfur-oxidizing chemolithoautotroph, was grown under microaerophilic conditions with thiosulfate as the only energy source. The rates of tetrathionate, thiosulfate, elemental sulfur (S^o) and sulfite oxidation were measured respirometrically with an oxygen electrode, using actively growing cells. Cells oxidized thiosulfate, elemental sulfur (S^o) and sulfite, but not tetrathionate. The thiosulfateoxidizing activity and elemental sulfur-oxidizing activity (SOA) were almost totally inhibited by 50 M myxothiazol (>80%), an inhibitor of the quinone-cytochrome b region, and by 10 M of the uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP) (>82%). Sulfite-oxidizing activity was also significantly inhibited (>60%) by 50 M myxothiazol and 10 M CCCP. 1 mM KCN totally inhibited (>90%) all respiratory activities. This study confirms that a sulfur-oxidizing activity appears during microaerophilic growth ofThiobacillus denitrificans strain RT on thiosulfate. The SOA is linked to the respiratory chain, probably releasing electrons in the quinone-cytochrome b region.To whom correspondence should be addressed. Submitted by R. Bachofen.     

Indirect visualization of hematopoietic colony-forming stem cell (CFUs) as a possible target cell forMycoplasma arthritidis

Summary Utilizing specific rabbit antiserum againstM. arthritidis together with complement, a portion of the marrow 7-day CFUs population of CBA mice infected with live mycoplasma organisms 1 day previously was shown to be inactivated. These cells might therefore be considered as candidate target cells forM. arthritidis. 11-day CFUs were unaffected by similar treatment.     

Antibacterial effect of 2-hydroxy-N-(3,4-dimethyl-5-isoxazolyl)-1,4-naphthoquinone-4-imine onStaphylococcus aureus

The mechanism by which a new naphthoquinone derivative, the 2-hydroxy-N-(3,4-dimethyl-5-isoxazolyl)-1,4-naphthoquinone-4-imine (INQI-E) has antibacterial effect againstStaphylococcus aureus was studied. The interaction of INQI-E with the bacteria was followed by absorption spectroscopy at 323 and 490 nm. The absorption band of INQI-E at 490 nm undergoes a hypochromic shift with a decrease of intensity. This effect was found to be reversible by oxygenation during the first hours of incubation. The participation of an oxidation-reduction process related to the respiratory chain was demonstrated by oxygen consumption. An increase in O₂ uptake and inhibition ofS. aureus growth was observed. Experiments with three inhibitors of the respiratory chain demonstrated that the pathway induced by INQI-E was antimycin-resistant and KCN- and salicylhydroxamic acid (SHAM)-sensitive, which suggests that INQI-E is capable of diverting the normal electron flow to an alternate superoxide-producing route. On the other hand, experiments with Tiron, a specific scavenger of superoxide, hindered the effect of INQI-E againstS. aureus, indicating that the inhibitory growth effect of this quinone-imine is mainly due to the production of the cytotoxic superoxide radical.     

The C4-dicarboxylate transport system ofRhizobium meliloti and its role in nitrogen fixation during symbiosis with alfalfa (Medicago sativa)

TheRhizobium meliloti C₄-dicarboxylate transport (Dct) system is essential for an effective symbiosis with alfalfa plants. C₄-dicarboxylates are the major carbon source taken up by bacteroids. Genetic analysis of Dct^– mutant strains led to the isolation of thedct carrier genedctA and the regulatory genesdctB anddctD. The carrier genedctA is regulated in free-living cells by the alternative sigma factor RpoN and the two-component regulatory system DctB/D. In addition, DctA is involved in its own regulation, possibly by interacting with DctB. In bacteroids, besides the DctB/DctD system an additional symbiotic activator is thought to be involved indctA expression. Further regulation ofdctA in the free-living state is reflected by diauxic growth of rhizobia, with succinate being the preferred carbon source. The tight coupling of C₄-dicarboxylate transport and nitrogen fixation is revealed by a reduced level of C₄-dicarboxylate transport in nitrogenase negative bacteroids.     

What leads from <Emphasis Type="Italic">dead-end?</Emphasis>

The 129 mouse strain develops congenital testicular germ cell tumors (TGCTs) at a low frequency. TGCTs in mice resemble the testicular tumors (teratomas) that occur in human infants. The genes that cause these tumors in 129 have not been identified. The defect at the Ter locus increases TGCT incidence such that 94% of 129-Ter/Ter males develop TGCTs. The primary effect of the Ter mutation is progressive loss of primordial germ cells (PGCs) during embryonic development. This results in sterility in adult Ter/Ter mice on all mouse strain backgrounds. However, on the 129 background, Ter causes tumor development in addition to sterility. Therefore, Ter acts as a modifier of 129-derived TGCT susceptibility genes. Ter was identified to be a mutation that inactivates the Dead-end1 (Dnd1) gene. In this perspective, I discuss the possible areas of future investigations to elucidate the mechanism of TGCT development due to Dnd1 inactivation. Received 29 September 2006; received after revision 29 January 2007; accepted 19 February 2007     

Die Wirkung energiereicher Strahlen auf die Atmungsfermentkette

Summary The irradiation sensitivity of the cytochrome system was investigated, using as examples baker's yeast andE. coli. A method is described with which the proportion of reduced cytochrome in cell suspensions could be measured spectrometrically. In contrast to isolated cytochrome, which is easily oxydized by Roentgen rays, the cytochrome of living cells is very stable to Roentgen rays. The electrone transport is not influenced by high energy rays. It is probable that the hydrogen transport is also undisturbed. Only if the cell is so damaged by high doses of rays that every exchange of substances is impossible, i.e. if the cell is dead, is the ferrocytochrome oxydized and perhaps also destroyed. The oxydation is thus a secondary process resulting from death of the cell by rays. It is supposed that the changes in energy metabolism described in the literature, especially the disturbances of oxydative phosphorylation, are also secondary processes.The concepts developed in radiochemistry on the action of high energy rays on organic compounds must therefore be especially tested in each case for their application in the living cell.     

Oenocyte and prothoracic gland activity inManduca sexta under varying photoperiod and light conditions

Summary Manduca sexta larvae were subjected to diapause-inducing and diapause-preventing photoperiods, using two types of fluorescents (Indorsun and Blacklight-blue). The oenocytes, prothoracic glands (PTG) and ecdysone levels were examined in 3-day-old 5th instar larvae, 2-day-old and 10-day-old pupae. Our results indicate that oenocytes and PTG cells tend to be more active under long photoperiods while oenocytes only are active under short photoperiods in pupae in diapause. UV light has a definite effect on oenocytes while PTG cells seem to be unaffected. Ecdysone and ecdysterone levels vary with PTG and oenocyte activity at the pupal stage. The significance of these findings is discussed.This work was supported by a research associateship to L.M. by the People's Republic of China and a grant from NSERC to B.J.R.P.     

Two classes of metabolites fromTheonella swinhoei are localized in distinct populations of bacterial symbionts

The marine spongeTheonella swinhoei (lithistid Family Theonellidae, Order Astrophorida) has yielded many important, bioactive natural products, most of which share structural features with bacterial natural products. The presence of microbial symbionts inT. swinhoei has been reported, and it was originally suggested that the cytotoxic macrolide swinholide A and many of the bioactive cyclic peptides fromT. swinhoei were all produced by symbiotic cyanobacteria. By transmission electron microscopy, we found four distinct cell populations to be consistently present inT. swinhoei: eukaryotic sponge cells, unicellular heterotrophic bacteria, unicellular cyanobacteria and filamentous heterotrophic bacteria. Purification and chemical analyses of each cell type showed the macrolide swinholide A to be limited to the mixed population of unicellular heterotrophic bacteria, and an anti-fungal cyclic peptide occurred only in the filamentous heterotrophic bacteria. Contrary to prior speculation, no major metabolites were located in the cyanobacteria or sponge cells.     

Periplasmic lysozyme inhibitor contributes to lysozyme resistance in <Emphasis Type="Italic">Escherichia coli</Emphasis>

The product of the Escherichia coli ORFan gene ykfE was recently shown to be a strong inhibitor of C-type lysozyme in vitro. The gene was correspondingly renamed ivy (inhibitor of vertebrate lysozyme), but its biological function in E. coli remains unknown. In this work, we investigated the role of Ivy in the resistance of E. coli to the bactericidal effect of lysozyme in the presence of outer-membrane-permeabilizing treatments. Both in the presence of lactoferrin (3.0 mg/ml) and under high hydrostatic pressure (250 MPa), the lysozyme resistance of E. coli MG1655 was decreased by knock-out of Ivy, and increased by overexpression of Ivy. However, knock-out of Ivy did not increase the lysozyme sensitivity of an E. coli MG1655 mutant previously described to be resistant to lysozyme under high pressure. These results indicate that Ivy is one of several factors that affect lysozyme resistance in E. coli, and suggest a possible function for Ivy as a host interaction factor in commensal and pathogenic E. coli.Received 12 February 2004; received after revision 11 March 2004; accepted 16 March 2004