Compartmentalisation of nitrogenase in a non-heterocystous cyanobacterium: Trichodesmium contortum

Abstract In Trichodesmium contortum , nitrogenase was detected in only a limited number (about 10%) of microscopically distinguishable, consecutively arranged cells in central regions of the trichomes. Cells with nitrogenase also contained the photosystem II associated pigment phycoerythrin. These cells were not distinguishable from other cells on a structural basis, but were clearly visible at low magnification microscopy as all in the zone were more compact and shorter than those on either side. The compartmentalisation of nitrogenase into a chain of cells and in a possibly photosynthetic environment represents a previously undescribed phenomenon. The nitrogenase containing cells apparently perform the O₂ protective function of heterocysts yet are different in several aspects.     

Generation of Mouse Small Intestinal Epithelial Cell Lines That Allow the Analysis of Specific Innate Immune Functions

Cell lines derived from the small intestine that reflect authentic properties of the originating intestinal epithelium are of high value for studies on mucosal immunology and host microbial homeostasis. A novel immortalization procedure was applied to generate continuously proliferating cell lines from murine E19 embryonic small intestinal tissue. The obtained cell lines form a tight and polarized epithelial cell layer, display characteristic tight junction, microvilli and surface protein expression and generate increasing transepithelial electrical resistance during in vitro culture. Significant up-regulation of Cxcl2 and Cxcl5 chemokine expression upon exposure to defined microbial innate immune stimuli and endogenous cytokines is observed. Cell lines were also generated from a transgenic interferon reporter (Mx2-Luciferase) mouse, allowing reporter technology-based quantification of the cellular response to type I and III interferon. Thus, the newly created cell lines mimic properties of the natural epithelium and can be used for diverse studies including testing of the absorption of drug candidates. The reproducibility of the method to create such cell lines from wild type and transgenic mice provides a new tool to study molecular and cellular processes of the epithelial barrier.     

Metabolic activation and toxicity of a DDT-metabolite, 3-methylsulphonyl-DDE, in the adrenal zona fasciculata in mice

Whole-body autoradiography of 14C-labelled 3-methylsulphonyl-DDE (3-MeSO2-DDE) in female C57BL mice revealed a heavy accumulation in the adrenal cortex. Fairly high radioactivity appeared in the nasal mucosa and fat, while the labelling of the liver was intermediate. The adrenal radioactivity remained largely unextracted in tissue-sections treated with organic solvents. In the liver and intestinal contents the radioactivity was partly extracted, whereas in all other tissues almost completely extracted. According to light microscopic autoradiography, the tissue-bound adrenal radioactivity was confined to the zona fasciculata, leaving the other adrenal zones devoid of bound material. Incubation of 3-MeSO2-DDE with adrenal tissue (300 X g supernatant) revealed a dose- and time-dependent covalent binding to protein and formation of water-soluble metabolites. The cytochrome P-450 inhibitors metyrapone and carbon monoxide inhibited both covalent binding and polar metabolite formation. Addition of reduced glutathione decreased binding, while polar metabolite formation was increased. Histopathological examination of adrenals from 3-MeSO2-DDE-treated mice revealed extensive vacuolation and necrosis of the zona fasciculata 1-12 days after single doses down to 25 mg/kg. Degenerative changes were observed at 12.5 mg/kg. In contrast to 3-MeSO2-DDE, 14C-labelled 3,3'-bis(methylsulphonyl)-DDE was not accumulated in the adrenal cortex. 3-MeSO2-DDE is thus a persistent environmental pollutant with a unique ability to produce acute toxicity subsequent to metabolic activation in a mammalian tissue.     

The effect of the 4B chromosomes of hexaploid wheat on the growth and regeneration of callus cultures

Summary Calli were initiated from immature embryos of nine lines of hexaploid wheat (Triticum aestivum L. em. Thell). These were the euploid lines Chinese Spring and Cappelle-Desprez, a line of Chinese Spring ditelocentric for the long arm of 4B, four substitution lines of Chinese Spring in which chromosome 4B has been replaced by its homologues from different wheat varieties and substituted into Chinese Spring and a substitution line of Besostaya I 4B into Cappelle-Desprez. The calli from these lines were found to differ in their growth rates and morphogenic and regenerative activities. The substitution of different 4B chromosomes into Chinese Spring significantly increased morphogenesis and shoot regeneration from callus. The potential for developing wheat lines with improved culture characteristics is discussed.     

The Nostoc-Nephroma symbiosis: localization, distribution pattern and levels of key proteins involved in nitrogen and carbon metabolism of the cyanobiont

The qualitative distribution and quantitative estimates of nitrogenase (EC 1.7.99.2), glutamine synthetase (EC 6.3.1.2), phycoerythrin and ribulose 1,5-bisphosphate carboxylase/oxygenase (EC 4.1.1.39) were studied in the cyanobacterium Nostoc residing in internal cephalodia of the tripartite lichen Nephroma arcticum L. Polyclonal antisera, raised in rabbit against the proteins, and goat anti-rabbit IgG conjugated to 10 nm gold were used as probes to detect the antigens by transmission electron microscopy. Western blot analyses demonstrated the monospecificity of the antisera. Nitrogenase was localized in heterocysts, with vegetative cells showing a label intensity comparable to the background. Distribution of the antigen within the heterocysts was uniform. Glutamine synthetase labelling was very low, but appeared to be distributed in both cell types. An intense phycoerythrin labelling was associated with the thylakoid region of the vegetative cells, whereas a much lower labelling was observed in the heterocyst. No significant differences were found between cyanobionts in younger and older cephalodia except for the nitrogenase labelling, which was higher in heterocysts of the cyanobiont in younger cephalodia. Most of the ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) label was present in vegetative cells. The Rubisco label was pronounced in the carboxysomes, whereas the label in the cytoplasm, on a unit area basis, was much lower. Heterocysts showed a label intensity similar to that of the vegetative cell cytoplasm. In Nostoc of the bipartite lichen Peltigera canina L., the Rubisco protein showed a comparable distribution pattern, but the average number of carboxysomes per vegetative cell was about 4 times higher.