A 718-kb DNA Sequence of the Escherichia coli K-12 Genome Corresponding to the 12.7-28.0 min Region on the Linkage Map

The 718,122 base pair (bp) sequence of the Escherichia coliK-12 genome corresponding to the region from 12.7 to 28.0 minuteson the genetic map is described. This region contains at least682 potential open reading frames, of which 278 (41%) have beenpreviously identified, 147 (22%) were homologous to other knowngenes, 138 (20%) are identical or similar to the hypotheticalgenes registered in databases, and the remaining 119 (17%) didnot show a significant similarity to any other gene. In thisregion, we assigned a cluster of cit genes encoding multienzymecitrate lyase, two clusters of fimbrial genes and a set of lysogenicphage genes encoding integrase, excisionase and repressor inthe e14 genetic element. In addition, a new valine tRNA gene,designated valZ, and a family of long directly repeated sequences,LDR-A, -B and -C, were found.     

The Effect of Potassium Nitrate on the Reduction of Phytophthora Stem Rot Disease of Soybeans, the Growth Rate and Zoospore Release of Phytophthora sojae

The effects of potassium nitrate (KNO₃) application on Phytophthora stem rot disease reduction of Glycine max (L.) Merr. cvs. Chusei-Hikarikuro and Sachiyutaka, and mycelium growth and zoospore release of a Phytophthora sojae isolate were investigated under laboratory conditions. The application of 4–30 m m KNO₃ prior to inoculation greatly reduced incidence of disease in the two soybean cultivars. Although a concentration of 20–30 m m KNO₃ led to a slight decrease in the growth rate of the PJ-H30 isolate on PDA medium, no significant relationship was observed between inhibition of the growth rate and disease reduction on application of 0.4–10 m m KNO₃. Disease suppression recorded in laboratory experiments using pathogen mycelium was due to the response of plant tissues rather than a direct inhibition of pathogen hyphal growth by the application of KNO₃. The extent of disease reduction was related to increased potassium concentration in plants of the two cultivars (except for some cases involving cv. Sachiyutaka), suggesting that differences existed between the two cultivars in terms of the effect of KNO₃ application on disease suppression. Scanning electron microscopic observation with fresh samples indicated marked accumulation of potassium at the penetration-stopping sites of P. sojae in the cortex layer of soybean plants treated with 30 m m KNO₃, compared with the non-treated control plants. The presence of 0.4–30 m m KNO₃ decreased the release of zoospores. These results suggest the possibility of applying a solution containing 20–30 m m of KNO₃ to decrease the incidence of disease in agricultural fields by the response of plant tissues to KNO₃.     

ADAMTS1 inhibits lymphangiogenesis by attenuating phosphorylation of the lymphatic endothelial cell-specific VEGF receptor

Angiogenesis and lymphangiogenesis play roles in malignant tumor progression, dissemination, and metastasis. ADAMTS1, a member of the matrix metalloproteinase family, is known to inhibit angiogenesis. Recombinant ADAMTS1 was shown to strongly inhibit angiogenesis. We investigated whether ADAMTS1 inhibited lymphangiogenesis in the present study. We examined cell proliferation and cell migration in normal human dermal lymphatic microvascular endothelial cells (HMVEC-dLy) transduced with or without adenoviral human ADAMTS1 gene therapy. We then examined the VEGFC/VEGFR3 signal transduction pathway in ADAMTS1-transduced HMVEC-dLy. Cell proliferation and tube formation in Matrigel were significantly lower with transduced ADAMTS1 than with control (non-transduced HMVEC-dLy). The phosphorylation of VEGFR3 was also attenuated by ADAMTS1 gene therapy in HMVEC-dLy. Immunoprecipitation assays revealed that ADAMTS1 formed a complex with VEGFC. Our results demonstrated that ADAMTS1 inhibited lymphangiogenesis in vitro. The data highlight the new function of ADAMTS1 in the regulation of lymphangiogenesis and the therapeutic potential of ADAMTS1 in cancer therapy.     

Structure–activity relationship study of glaziovianin A against cell cycle progression and spindle formation of HeLa S3 cells

Various derivatives of glaziovianin A, an antitumor isoflavone, were synthesized, and the cytotoxicity of each against HeLa S₃ cells was investigated. Compared to glaziovianin A, the O⁷-allyl derivative was found to be more cytotoxic against HeLa S₃ cells and a more potent M-phase inhibitor.     

Quantitative analysis of the redox states of cytochromes in a living L929 (NCTC) cell by resonance Raman microspectroscopy

Raman spectra and images of a living L929 (NCTC) cell have been measured with 532 nm excitation. Both reduced and oxidized forms of cytochromes b and c (cyt b and cyt c) have been observed in situ without any pretreatment. The redox states of cyts b and c have been assessed quantitatively with a spectral analysis. It has been found that reduced cyt c is more abundant than oxidized cyt c, while oxidized cyt b is slightly more abundant than reduced cyt b in a living cell. (© 2013 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

A complete enzymatic capacity for biosynthesis of docosahexaenoic acid (DHA, 22 : 6n–3) exists in the marine Harpacticoida copepod Tigriopus californicus

The long-standing paradigm establishing that global production of Omega-3 (n–3) long-chain polyunsaturated fatty acids (LC-PUFA) derived almost exclusively from marine single-cell organisms, was recently challenged by the discovery that multiple invertebrates possess methyl-end (or ωx) desaturases, critical enzymes enabling the biosynthesis of n–3 LC-PUFA. However, the question of whether animals with ωx desaturases have complete n–3 LC-PUFA biosynthetic pathways and hence can contribute to the production of these compounds in marine ecosystems remained unanswered. In the present study, we investigated the complete enzymatic complement involved in the n–3 LC-PUFA biosynthesis in Tigriopus californicus, an intertidal harpacticoid copepod. A total of two ωx desaturases, five front-end desaturases and six fatty acyl elongases were successfully isolated and functionally characterized. The T. californicus ωx desaturases enable the de novo biosynthesis of C₁₈ PUFA such as linoleic and α-linolenic acids, as well as several n–3 LC-PUFA from n–6 substrates. Functions demonstrated in front-end desaturases and fatty acyl elongases unveiled various routes through which T. californicus can biosynthesize the physiologically important arachidonic and eicosapentaenoic acids. Moreover, T. californicus possess a Δ4 desaturase, enabling the biosynthesis of docosahexaenoic acid via the ‘Δ4 pathway’. In conclusion, harpacticoid copepods such as T. californicus have complete n–3 LC-PUFA biosynthetic pathways and such capacity illustrates major roles of these invertebrates in the provision of essential fatty acids to upper trophic levels.     

Surfactant Uptake Dynamics in Mammalian Cells Elucidated with Quantitative Coherent Anti-Stokes Raman Scattering Microspectroscopy

The mechanism of surfactant-induced cell lysis has been studied with quantitative coherent anti-Stokes Raman scattering (CARS) microspectroscopy. The dynamics of surfactant molecules as well as intracellular biomolecules in living Chinese Hamster Lung (CHL) cells has been examined for a low surfactant concentration (0.01 w%). By using an isotope labeled surfactant having CD bonds, surfactant uptake dynamics in living cells has been traced in detail. The simultaneous CARS imaging of the cell itself and the internalized surfactant has shown that the surfactant molecules is first accumulated inside a CHL cell followed by a sudden leak of cytosolic components such as proteins to the outside of the cell. This finding indicates that surfactant uptake occurs prior to the cell lysis, contrary to what has been believed: surface adsorption of surfactant molecules has been thought to occur first with subsequent disruption of cell membranes. Quantitative CARS microspectroscopy enables us to determine the molecular concentration of the surfactant molecules accumulated in a cell. We have also investigated the effect of a drug, nocodazole, on the surfactant uptake dynamics. As a result of the inhibition of tubulin polymerization by nocodazole, the surfactant uptake rate is significantly lowered. This fact suggests that intracellular membrane trafficking contributes to the surfactant uptake mechanism.     

Influence of Asian Dust Particles on Immune Adjuvant Effects and Airway Inflammation in Asthma Model Mice

Objective

An Asian dust storm (ADS) contains airborne particles that affect conditions such as asthma, but the mechanism of exacerbation is unclear. The objective of this study was to compare immune adjuvant effects and airway inflammation induced by airborne particles collected on ADS days and the original ADS soil (CJ-1 soil) in asthma model mice.

Methods

Airborne particles were collected on ADS days in western Japan. NC/Nga mice were co-sensitized by intranasal instillation with ADS airborne particles and/or Dermatophagoides farinae (Df), and with CJ-1 soil and/or Df for 5 consecutive days. Df-sensitized mice were stimulated with Df challenge intranasally at 7 days after the last Df sensitization. At 24 hours after challenge, serum allergen specific antibody, differential leukocyte count and inflammatory cytokines in bronchoalveolar lavage fluid (BALF) were measured, and airway inflammation was examined histopathologically.

Results

Co-sensitization with ADS airborne particles and Df increased the neutrophil and eosinophil counts in BALF. Augmentation of airway inflammation was also observed in peribronchiolar and perivascular lung areas. Df-specific serum IgE was significantly elevated by ADS airborne particles, but not by CJ-1 soil. Levels of interleukin (IL)-5, IL-13, IL-6, and macrophage inflammatory protein-2 were higher in BALF in mice treated with ADS airborne particles.

Conclusion

These results suggest that substances attached to ADS airborne particles that are not in the original ADS soil may play important roles in immune adjuvant effects and airway inflammation.