Control of Nitrogenase mRNA Levels by Products of Nitrate Assimilation in the Cyanobacterium Anabaena sp. Strain PCC 7120

Nitrate inhibited nitrogenase synthesis and heterocyst development in the cyanobacterium Anabaena sp. strain PCC 7120. Inhibition of dinitrogen fixation by nitrate did not take place, however, in nitrate reductase-deficient derivatives of this strain. Hybridization of total RNA isolated from cells grown on different nitrogen sources with an internal fragment of the nifD gene showed that regulation of nitrogenase activity by nitrate is exerted through a negative control of the nitrogenase mRNA levels.     

Restoration of normal stamen development and pollen formation by fusion of different cytoplasmic male-sterile cultivars of Nicotiana tabacum

Summary Fusion of two cytoplasmic male-sterile cultivars of Nicotiana tabacum, one with N. bigelovii cytoplasm and one with N. undulata cytoplasm, resulted in the restoration of male fertility in cybrid plants. All male-fertile cybrids exhibited fused corollas, which is characteristic for the cultivar with N. undulata cytoplasm, while their stamen structures varied from cybrid to cybrid, some producing stamens with anthers fused to petal-like appendages and one producing stamens of a normal appearance for N. tabacum. Restriction enzyme digestion and agarose gel electrophoresis of mitochondrial DNA showed that mitochondrial DNA of the fertile cybrids was more similar to the male-sterile cultivar with the cytoplasm of N. undulata than to the cultivar with N. bigelovii cytoplasm. Some restriction fragments were unique to the male-fertile cybrids. Comparisons between stamen structure and mitochondrial DNA for eight fertile progeny from one cybrid plant led to the identification of several restriction fragments that appeared at enhanced levels in connection with normal stamen development.     

Clomipramine actions on firing rate in septal nuclei of the rat are not related to anaesthesia (urethane).

An increased firing rate in lateral septal nuclei (LSN) appears in urethane-anesthetized rats after several acute drug and non-drug human antidepressant treatments. A still more pronounced increase in firing rate is produced in LSN after clomipramine (CMI) long-term treatment. In spite of urethane is a widely used anesthetic for single unit extracellular recordings, it modifies evoked potentials wave-form. Therefore, present study discards urethane interaction with CMI in LSN single unit extracellular recordings. CMI was acutely injected (1.25 mg/kg: IP) either to urethane-anesthetized, or non-anesthetized encephale-isolé rats. The CMI treated groups showed higher rates of firing in LSN regardless of the use of general anesthesia during recordings. Another group of urethane-anesthetized rats received intracerebroventricular (ICV) microinjections of CMI (100 micrograms/10 microliters/1 min). An amount of 42.8% of LSN-recorded neurons responded with a long-lasting increased firing rate. Results discard urethane and CMI interactions. Additionally, systemic actions of CMI on firing rate of LSN are reproduced by ICV/route microinjections.     

High-level expression of a tobacco chitinase gene in Nicotiana sylvestris. Susceptibility of transgenic plants to Cercospora nicotianae infection

Endochitinases (E.C. 3.2.14, chitinase) are believed to be important in the biochemical defense of plants against chitin-containing fungal pathogens. We introduced a gene for class I (basic) tobacco chitinase regulated by Cauliflower Mosaic Virus 35S-RNA expression signals into Nicotiana sylvestris. The gene was expressed to give mature, enzymatically active chitinase targeted to the intracellular compartment of leaves. Most transformants accumulated extremely high levels of chitinase-up to 120-fold that of non-transformed plants in comparable tissues. Unexpectedly, some transformants exhibited chitinase levels lower than in non-transformed plants suggesting that the transgene inhibited expression of the homologous host gene. Progeny tests indicate this effect is not permanent. High levels of chitinase in transformants did not substantially increase resistance to the chitin-containing fungus Cercospora nicotiana, which causes Frog Eye disease. Therefore class I chitinase does not appear to be the limiting factor in the defense reaction to this pathogen.     

Identification, genetic analysis and characterization of a sugar-non-specific nuclease from the cyanobacterium Anabaena sp. PCC 7120

A nuclease that could be recovered from the supernatant of cultures, as well as from cell-free extracts, of the cyanobacterium Anabaena sp. PCC 7120 was identified as a 29 kDa polypeptide by its ability to degrade DNA after electrophoresis in DNA-containing SDS-polyacrylamide gels. Some clones of a gene library of strain PCC 7120 established in Escherichia coli were found to produce the 29 kDa nuclease. The nucA gene encoding this nuclease was subcloned and sequenced. The deduced polypeptide, NucA, had a molecular weight of 29,650, presented a presumptive signal peptide in its N-terminal region and showed homology to the products of the nuc gene from Serratia marcescens and the NUC1 gene from Saccharomyces cerevisiae. The NucA protein from Anabaena itself, or from the cloned nucA gene expressed in E. coli, catalysed the degradation of both RNA and DNA, had the potential to act as an endonuclease, and functioned best in the presence of Mn2+ or Mg2+. An Anabaena nucA insertional mutant was generated which failed to produce the 29 kDa nuclease.     

gltF, a member of the gltBDF operon of Escherichia coli, is involved in nitrogen-regulated gene expression

We report here the construction and analysis of insertional mutations in each of the three genes of the gltBDF operon and the nucleotide sequence of the region downstream from gltD. Two open reading frames were identified, the first of which corresponds to gltF. The gltB and gltD genes code for the large and small subunits, respectively, of the enzyme glutamate synthase (GOGAT). gltF codes for a protein, with a molecular mass of 26,350 Da, which is required for Ntr induction. Histidase synthesis was determined as a measure of Ntr function. First, insertions in gltB, gltD or gltF all prevent Ntr induction. Second, complementation analysis indicates that high-level expression of both the gltD and gltF genes is required for the induction of the Ntr enzymes under nitrogen-limiting conditions, indicating that the phenotype of the gltB insertion probably results from polarity on gltD and gltF. Third, glutamate-dependent repression of the glt operon appears to be mediated by the product of the gltF gene. Thus, the gltBDF operon of Escherichia coli is involved in induction of the so-called Ntr enzymes in response to nitrogen deprivation, as well as in glutamate biosynthesis.     

Glomerular lesions in Aleutian disease of mink (Mustela vison): a morphological and differential morphometrical study.

A morphological and morphometrical study has been carried out on glomerular lesions in mink with spontaneous Aleutian disease, using the WHO classification for Systemic Lupus Erytematous Nefritis. 154 renal samples from sick animals and 10 samples from uninfected mink were processed by routine histopathological techniques and metacrylate inclusions. The samples were studied quantitatively with an automatic image analyzer. 5 forms of glomerulonephritis (GN) were identified: mesangial glomerulonephritis (n = 13), focal and segmental GN (n = 10), diffuse GN (n = 99), membranous GN (n = 12) and advanced sclerosing GN (n = 10) and were associated with the degree of interstitial plasmocytosis. Glomerule morphometry was shown to be an excellent method for identifying the type of lesion, while it quantified the participation of various glomerular elements in the lesion.     

Stimulation of intestinal adenylate cyclase by cholera toxin in malnourished rats

The stimulation of intestinal adenylate cyclase by cholera toxin (CT) was studied in normal and malnourished rats 4 to 24 hr after a 30-min incubation of intestinal loops with the toxin. Whereas in control rats the enzyme activity returned to basal levels after 12 hr of incubation, in malnourished rats the activity of the enzyme remained significantly elevated even after 24 hr of the initial incubation. Malnourished animals had a reduced turnover rate of intestinal cells as determined by thymidine kinase activity. The delayed turnover of intoxicated cells may account for continuous activation of mucosal adenylate cyclase and possibly for prolongation of diarrhea in malnutrition.     

Wheat germ agglutinin-resistant variant of EL4 containing altered oligosaccharides as a target cell for cytotoxic T cells

A lectin-resistant variant of the murine EL4 lymphocytic leukemia cell line was selected in the presence of wheat germ agglutinin for low levels of cell-surface sialic acid. H-2Kb was the major internally radiolabeled H-2b molecule on the cell-surface of WD1, and it was not sialylated, as determined by two-dimensional gel analysis. Endo-beta-N-acetylglucosaminidase H treatment of the WD1 membrane fractions suggested that the oligosaccharides on the cell-surface H-2Kb molecule were complex, but nonsialylated. Monoclonal antibody inhibition of the allogeneically primed cell-mediated cytotoxicity (CMC) reaction indicated that the T cells (BALB/c anti-EL4; H-2d anti-H-2b) were specific only for the H-2Kb target cell antigen. These WD1 variant cells were used as targets in the CMC assay using anti-H-2Kb T cells and compared with the parent EL4 in vitro line. The change in the cell-surface oligosaccharide did not affect the susceptibility to lysis by the cytotoxic T lymphocytes even though there were 2.5-fold more H-2Kb antigens on the WD1 variant cell (1.5 X 10(5) sites/cell) than on the parent EL4 in vitro cell (5.9 X 10(4) sites/cell). It was possible to isolate highly purified preparations of H-2Kb from either the EL4 or the WD1 line using a monoclonal antibody affinity column. Interestingly, the variant WD1 cell would no longer grow in the peritoneal cavity of the syngeneic C57BL/6 mouse.