Transformation of colonial Escherichia coli on solid media

We report that colonial Escherichia coli cells on various solid media can develop modest genetic competence. Using an on-filter culture system, we found that E. coli colonies on CaCl2-containing agar were transformed in the presence of plasmid DNA. Interestingly, transformation also occurred on LB-agar, various moist foods and even on H2O-agar. These results suggest that some populations of colonial E. coli in various environments could become transformable regardless of the surrounding Ca2+ concentration.     

Morphogenic protein epimorphin protects intestinal epithelial cells from oxidative stress by the activation of EGF receptor and MEK/ERK, PI3 kinase/Akt signals

Epimorphin is a mesenchymal protein that regulates morphogenesis of epithelial cells. Our preliminary study suggested a novel function of epimorphin in enhancing survival of intestinal epithelial cells (IEC). Oxidative stress leads to cell injury and death and is suggested to be a key contributor to pathogenesis of inflammatory bowel disease. This study was conducted to determine whether epimorphin protects IEC from oxidative stress. Rat intestinal epithelial cell line IEC-6 was cultured with epimorphin (10 and 20 mug/ml), and the life span of IEC was assessed. The mean life span of IEC-6 cells was prolonged 1.9-fold (P < 0.0006) by treatment with epimorphin. We then examined the epimorphin signaling pathways. Epimorphin phosphorylated epidermal growth factor (EGF) receptor, activated the MEK/extracellular signal-regulated kinase 1/2 mitogen-activated protein kinase and phosphatidylinositol 3 (PI3) kinase/Akt pathways, phosphorylated Bad, and induced Bcl-X(L) and survivin. Hydrogen peroxide (1 mM) induced cell death in 92% of IEC-6 cells, but epimorphin dramatically diminished (88.7%) cell death induced by hydrogen peroxide (P < 0.0001). This protective effect of epimorphin was significantly attenuated by inhibitors of MEK and PI3 kinase (P < 0.0001) or EGF receptor-neutralizing antibody (P = 0.0007). In wound assays, the number of migrated cells in the wound area decreased (72.5%) by treatment with 30 muM hydrogen peroxide, but epimorphin increased the number of migrated cells 3.18-fold (P < 0.0001). These results support a novel function of epimorphin in protecting IEC from oxidative stress. This anti-oxidative function of epimorphin is dramatic and is likely mediated by the activation of EGF receptors and the MEK/extracellular signal-regulated kinase and PI3 kinase/Akt signaling pathways and through the induction of anti-apoptotic factors.     

Generation of Superoxide Anion and Localization of CuZn-Superoxide Dismutase in the Vascular Tissue of Spinach Hypocotyls: Their Association with Lignification

The sites of generations of superoxide anions and hydrogen peroxidein cross sections of hypocotyls from spinach seedlings werelocated by staining with nitroblue tetrazolium (NBT) and withstarch-iodide, respectively. Formazan, produced upon the reductionof NBT by superoxide, was observed mainly in the vascular tissueonly in the presence of inhibitors of CuZn-superoxide dismutase(CuZn-SOD), and its formation was suppressed under anaerobicconditions. Thus, NBT was reduced to formazan specifically bythe superoxide anions generated in vascular tissue. The reductionof NBT was suppressed by inhibitors of NAD(P)H oxidase, butneither by cyanide nor azide, indicating the involvement ofNAD(P)H oxidase in the generation of superoxide anions in thevascular tissue. Starch-I₂ complex also was formed in the vasculartissue, but not in the presence of either the CuZn-SOD inhibitoror the NAD(P)H oxidase inhibitor, indicating that the hydrogenperoxide is produced via the catalytic disproportionation withCuZn-SOD of the superoxide generated by NAD(P)H oxidase. Generationsof superoxide anions and hydrogen peroxide in the vascular tissuewere particularly apparent in the xylem and associated withthe sites of distribution of CuZn-SOD as determined by an immunohistochemicalmethod, and also with the location of lignin as determined bythe phloroglucin-HCI reaction. ⁴Present address: Chemical Research Laboratories, Toray IndustriesInc., 9–1 Oe-cho, Minato-ku, Ngagoya, 455 Japan     

Experimental Myocarditis Induced in Mice by Infection with Influenza A2 Virus

A mouse model was established for the study of acute myocarditis that occurs during influenza infection. Challenge with more than 10 LD₅₀ of mouse-adapted influenza A₂ virus (H₂N₂) induced myocarditis macroscopically discernible as white, irregularly shaped lesions which were shown by histological examination to consist of necrotic myofibers surrounded by infiltrating mononuclear inflammatory cells. After challenge with 10 LD₅₀ of the virus, macroscopic myocarditis was found to advance in a progressive manner up to the 7th day, while the virus titer in the heart reached its peak on the 2nd day and began to decrease on the 5th day of infection. However, development of myocarditis was significantly suppressed in mice which were irradiated with 400 R of X-rays before infection. In addition, myocarditis did not develop in congenitally athymic nude mice. These data indicate that myocarditis was not brought about by viral action directly, but that it was mediated by some function of the host against viral in-vasion, which was abolished by X-irradiation. The data also suggest that T cells played a key role in the development of myocarditis.     

Glycerol production from sugars with phosphoglycerate mutasedeficientSaccharomyces cerevisiae partially resistant to glucose repression

Summary Mutants partially resistant to the repressive effect of glucose have been isolated from aSaccharomyces cerevisiae strain totally deficient in phosphoglycerate mutase activity (EC 5.4.2.1) by a selection procedure involving the catabolite-repressive effect of 5-thio-d-glucose (5TG). These mutants are able to resist glucose concentrations up to 15 g L^–1 and exhibit several non-repressed metabolic pathways such as gluconeogenesis, glyoxylic shunt or mitochondrial respiratory chain. Moreover, when these mutants are grown in aerobiosis on ethanol and glucose as sole substrates, glucose is mainly converted into glycerol in order to maintain a normal redox balance. Optimal glucose and oxygen concentrations have been defined for resting cells in order to obtain a glycerol yield from glucose close to 100%. The physiological characteristics of one of these mutants led us to consider an application of this yeast strain in reducing the ethanol content of wines previously lowered in ethanol content by physical processes.     

In vitro induction of stigma-like and style-like structures from receptacle tissue in Nicotiana tabacum L.

Stigma-like and style-like structures were induced from the receptacle tissue in tobacco (Nicotiana tabacum L.). The presence of kinetin was necessary to induce these structures. The structures have a form similar to and the same function as normal stigmas and styles.