On herbicidal actions

A theoretical interpretation of the herbicidal action of the 2,4-Dichlorophenoxyacetic acid is suggested.     

Gaseous NO2 as a regulator for ammonia oxidation of Nitrosomonas eutropha

Cells of Nitrosomonas eutropha strain N904 that were denitrifying under anoxic conditions with hydrogen as electron donor and nitrite as electron acceptor were unable to utilize ammonium (ammonia) as an energy source. The recovery of ammonia oxidation activity was dependent on the presence of NO₂. Anaerobic ammonia oxidation activity was observed in a helium atmosphere supplemented with 25 ppm NO₂ after 20 h. Ammonia oxidation activity was detected after 2–3 days using an oxic atmosphere with 25 ppm NO₂. In contrast, ammonia consumption started after 8–9 days under oxic conditions without the addition of NO₂; in this case, small amounts of NO and NO₂ were detected and their concentrations increased with increasing ammonia oxidation activities. Hardly any ammonia oxidation was detected when nitrogen oxides were removed by intensive aeration. It would seem, therefore, that NO₂ is the master regulatory signal for ammonia oxidation in Nitrosomonas eutropha. Anaerobic ammonia oxidation activity was inhibited by the addition of NO. This inhibition was partly compensated by either increasing the NO₂ concentration or by using 2,3-dimercapto-1-propane-sulfonic acid as a NO binding substrate. DMPS was inhibitory to nitrification under oxic conditions, while increased amounts of NO or NO₂ led to increased oxidation activities.     

Reptilian melanomacrophages function under conditions of hypothermia: observations on phagocytic behavior

Melanomacrophages (MMs) were removed from livers of turtles of three North American families and cultured. J774 mammalian macrophages were similarly cultured and the MMs were exposed to E. coli(fluor) at approximately 2, 7, 27, and 37 degrees C. At least one third of the MMs continued to function at the low temperatures where less than 2% of the mammalian cells incorporated bacteria deeply into the cytoplasm. In most instances, when the bacteria were not internalized deeply into the cytoplasm, they became stationary just inside, or within, the cell membrane. The MMs were significantly less efficient than the mammalian cells at 37 degrees C and significantly more efficient at 2 and 7 degrees C. In general, it appears that MMs are never as efficient as mammalian macrophages under the most ideal temperatures for the cell but they are capable of functioning at reasonable levels at temperature extremes. The observations are suggestive of a genetic mechanism functioning in the MMs that is rarely expressed in J774 cells under conditions of hypothermia. MMs in vitro and probably in vivo consume bacteria, fungi, attach to helminth eggs, and consume old erythrocytes.     

Use of the human EF-1alpha promoter for expression can significantly increase success in establishing stable cell lines with consistent expression: a study using the tetracycline-inducible system in human cancer cells

Establishing cells with an exogenously introduced gene of interest under the inducible control of tetracycline (Tc) initially requires clonal cell lines stably expressing the tetracycline activator (tTA or rtTA). The originally described plasmid vectors expressing tTA/rtTA are driven by the cytomegalovirus (CMV) immediate early (IE) promoter-enhancer, known for its robust activity in a wide spectrum of cell types. While many reports testify to the utility and efficacy of this construct, instances of inexplicable failure to establish cell lines having inducible expression of the cDNA under study are encountered. Spontaneous extinction of CMV promoter activity in cells has been observed in a temporal and cell type-dependent manner. This could be a contributing factor in the failure to establish Tc-responsive cell lines. We here report that a change of the expression cassette to the human elongation factor-1alpha (EF-1alpha) promoter has permitted successful establishment of several inducible cell lines from diverse human tumor tissue origins. We interpret these results to imply that extinction of rtTA (or tTA) expression might be a significant factor in the lack of success in establishing Tc-inducible cell lines. Moreover, the present findings have general relevance to experiments requiring the use of stable cell lines.     

Functional domains of the ubiquitous chromatin protein DEK

DEK was originally described as a proto-oncogene protein and is now known to be a major component of metazoan chromatin. DEK is able to modify the structure of DNA by introducing supercoils. In order to find interaction partners and functional domains of DEK, we performed yeast two-hybrid screens and mutational analyses. Two-hybrid screening yielded C-terminal fragments of DEK, suggesting that DEK is able to multimerize. We could localize the domain to amino acids 270 to 350 and show that multimerization is dependent on phosphorylation by CK2 kinase in vitro. We also found two DNA binding domains of DEK, one on a fragment including amino acids 87 to 187 and containing the SAF-box DNA binding motif, which is located between amino acids 149 and 187. This region is sufficient to introduce supercoils into DNA. The second DNA binding domain is located between amino acids 270 and 350 and thus overlaps the multimerization domain. We show that the two DNA-interacting domains differ in their binding properties and in their abilities to respond to CK2 phosphorylation.     

Diversity of coexisting <Emphasis Type="Italic">Planktothrix</Emphasis> (Cyanobacteria) chemotypes deduced by mass spectral analysis of microystins and other oligopeptides

Cyanobacteria are reported to produce secondary metabolites of which toxic and bioactive peptides are of scientific and public interest. Many peptides are synthesized by the non-ribosomal peptide synthesis pathway and their presence is a stable feature of individual clones. We isolated 18 clonal strains of Planktothrix from a single water sample from lake Maxsee near Berlin and analyzed them by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, HPLC, and PCR for their production of peptides and the presence of microcystin synthetase genes. Microcystins could be detected in seven of the strains with considerable variability of contents and numbers of structural variants. Other known peptides like anabaenopeptins B and E/F, microviridin I, and prenylagaramide B and new variants of known peptide classes like aeruginosins and cyanopeptolins were detected in some strains while lacking in others. The 18 strains represented 15 chemotypes with respect to their peptide patterns. In contrast, all strains were morphologically very similar with respect to cell dimensions and pigmentation. Given the diversity of chemotypes among the randomly selected isolates, an immense diversity of chemotypes in the entire population can be assumed.     

Sequential dimerization of human zipcode-binding protein IMP1 on RNA: a cooperative mechanism providing RNP stability

Active cytoplasmic RNA localization depends on the attachment of RNA-binding proteins that dictate the destination of the RNA molecule. In this study, we used an electrophoretic mobility-shift assay in combination with equilibrium and kinetic analyses to characterize the assembly of the human zipcode-binding protein IMP1 on targets in the 3′-UTR from Igf-II mRNA and in H19 RNA. In both cases, two molecules of IMP1 bound to RNA by a sequential, cooperative mechanism, characterized by an initial fast step, followed by a slow second step. The first step created an obligatory assembly intermediate of low stability, whereas the second step was the discriminatory event that converted a putative RNA target into a ‘locked’ stable RNP. The ability to dimerize was also observed between members of the IMP family of zipcode-binding proteins, providing a multitude of further interaction possibilities within RNP granules and with the localization apparatus.