Synthesis of (1→2)-β-d-Glucan by Cell-free Extracts ofAgrobacterium radiobacter IFO 12665b 1 andRhizobium phaseoli AHU 1133

Particulate preparations from Agrobacterium radiobacter IFO 12665b 1 and Rhizobium phaseoli AHU 1133 have been shown to catalyze the synthesis of (1→2)-β-d-glucan from UDP-d-[¹⁴C]glucose. The (1→2)-β-d-glucans synthesized are suggested to be in a cyclic form without other glycosidic linkages and to consist of a mixture of several components with degrees of polymerization of 17 and more. The enzyme systems from A. radiobacter IFO 12665b 1 and R. phaseoli AHU 1133 both required Mn²⁺ and had optimum activities at pH 7.5 ~ 8, and their Km values for UDP-d-[¹⁴C]glucose were 5 × 10~⁵ m and 3.3 × 10^?5 m, respectively.     

Development of biological soil disinfestations in Japan

Biological soil disinfestations (BSDs) were developed separately in Japan and in The Netherlands as an alternative to chemical fumigations. In Japan, it was developed based on the knowledge of irrigated paddy rice and upland crop rotation system that was rather tolerant of soil-borne disease development. The methods consist of application of easily decomposable organic matter, irrigation, and covering the soil surface with plastic film, thereby inducing anaerobic (reductive) soil conditions and suppressing many soil-borne pests including fungi, bacteria, nematodes, and weeds. The methods are widely used by organic farmers in the area where residences and agricultural fields are intermingled. To note one advantage of these methods, maintenance of soil suppressiveness to Fusarium wilt of tomato was suggested, while soil treated with chloropicrin became conducive to the disease. Suppression of soil-borne fungal pathogens by BSDs might be attributed to anaerobicity and high temperature, organic acids generated, and metal ions released into soil water. Contributions of respective factors to suppression of respective pathogens might be diverse. Presumably, these factors might vary on the fungal community structure in BSD-treated soil. These factors also work in paddy fields. Therefore, the BSDs developed in Japan are probably a method to raise the efficacy of paddy–upland rotation through intensive organic matter application and through maintenance of a strongly anaerobic (reductive) soil condition.     

(+)-Abscisic Acid and Two Compounds Showing Chlorophyll Degradation Activity in Cuscuta pentagona Engelm

The purified polyethylene glycol (PEG) dehydrogenase from cells of a synergistic mixed culture of Flavobacterium and Pseudomonas species showed a similar absorption spectrum to those of other quinoproteins reported so far. The prosthetic group of the PEG dehydrogenase after extraction with cold methanol and purification by DEAE-Sephadex A-25 column chromatography and Sephadex G-25 gel filtration showed the same elution profiles as those of authentic pyrrolo-quinoline quinone (PQQ). Absorption and fluorescence spectra of the purified prosthetic group and its prosthetic group capability for glucose dehydrogenase indicated that it was identical with authentic PQQ.

The enzyme was induced during bacterial cell growth on a medium containing PEG 6000 as a sole source of carbon. The purified enzyme oxidized primary alcohols of C₂-C₁₆ and the corresponding aldehydes of C₄-C₇. The enzyme also reacted with nonionic surfactants containing PEG residues. The enzyme reduced 2,6-dichlorophenolindophenol (DCIP) and the Km value for DCIP was calculated to be 1.4 × 10^?4m. The DCIP reductase activity was inhibited by carbonyl reagents like semicarbazide, hydrazine, hydroxylamine and 1,4-benzoquinone. 1,4-Benzoquinone inhibited the DCIP reductase activity competitively as to DCIP.     

Arabidopsis thaliana MAP65-1 and MAP65-2 function redundantly with MAP65-3/PLEIADE in cytokinesis downstream of MPK4

Plant cytokinesis occurs by the growth of cell plates from the interior to the periphery of the cell. These dynamic events in cytokinesis are mediated by a plant-specific microtubule (MT) array called the phragmoplast, which consists of bundled antiparallel MTs between the two daughter nuclei. The NACK-PQR pathway, a NACK1 kinesin-like protein and mitogen activated protein kinase (MAPK) cascade, is a key regulator of plant cytokinesis through the regulation of phragmoplast MTs. The MT-associated protein MAP65 has been identified as one of the structural components of MT assays involved in cell division, and we recently showed that Arabidopsis AtMAP65-3/PLEIADE (PLE) is a substrate of MPK4 that is a component of the NACK-PQR pathway in Arabidopsis. Here we show that AtMAP65-1 and AtMAP65-2 are also phosphorylated by MPK4. AtMAP65-1 and AtMAP65-2 that localize to the phragmoplast were phosphorylated by MPK4 in vitro. Although mutants of the Arabidopsis AtMAP65-1 and AtMAP65-2 genes exhibited a wild-type phenotype, double mutations of AtMAP65-3 and AtMAP65-1 or AtMAP65-2 caused more severe growth and cytokinetic defects than the single atmap65-3/ple mutation. These results suggest that AtMAP65-1 and AtMAP65-2 also function in cytokinesis downstream of MPK4.Key words: MAP65, microtubule-associated protein, MAPK, cytokinesis, phragmoplast, microtubule, arabidopsisMitogen-activated protein kinase (MAPK) cascades are highly conserved signaling modules in eukaryotes, and are involved in various signaling processes in plant development, cell division and responses to endogenous or exogenous stimuli.¹ The NACK-PQR pathway, one of the best-characterized MAPK cascades in plants, has been identified as a key regulator of plant cytokinesis in tobacco. This pathway is composed of NPK1 MAPK kinase kinase (MAPKKK), NQK1/NtMEK1 MAPK kinase (MAPKK), NRK1/NTF6 MAPK and NACK1 kinesin-like protein, an activator of NPK1 MAPKKK.^2–5 During cytokinesis, all these components are localized on the equator of the phragmoplast, which is the plant-specific cytokinetic apparatus organized by microtubules (MTs). Downstream of this pathway, tobacco MAP65-1, an MT-associated protein, is phosphorylated by NRK1/NTF6 MAPK and phosphorylated MAP65-1 is localized to the equator of the phragmoplast.⁶ Phosphorylation of MAP65-1 by NRK1/NTF6 decreases the ability of MAP65-1 to bundle MTs, suggesting that the NACK-PQR pathway regulates expansion of the phragmoplast through the phosphorylation of MAP65.⁶The NACK-PQR pathway also seems to be conserved in Arabidopsis and rice. Several orthologs of components of the NACK-PQR pathway except for MAPK have been identified independently as regulators of cytokinesis in these plants.^3,5,7–14 Recently we reported that ANP MAPKKKs, MPK6/ANQ MAPKK and MPK4 MAPK biochemically constitute the MAPK pathway and HINKEL/AtNACK1 functions as an activator of ANP MAPKKKs.¹⁵ In addition, we revealed that MPK4 MAPK is localized to cell plates during cytokinesis, is required for cytokinesis in Arabidopsis and phosphorylates AtMAP65-3.¹⁶ Although AtMAP65-3 is proposed to be involved in cytokinesis,^17,18 and AtMAP65-1 is supposed to be a substrate of MPK4 based on a series of experiments,^6,19,20 the involvement in cytokinesis of other closely related members of the Arabidopsis MAP65 family, AtMAP65-1 and AtNAP65-2, has yet to be tested. In this report, we suggest redundant functions of these MAP65 molecules in cytokinesis of Arabidopsis.     

Interferon-α/β and anti-fibroblast growth factor receptor 1 monoclonal antibody suppress hepatic cancer cells in vitro and in vivo

Background

Hepatocellular carcinoma (HCC) is the most commonly occurring primary liver cancer and ranks as the fifth most frequently occurring cancer, overall, and the third leading cause of cancer deaths, worldwide. At present, effective therapeutic options available for HCC are limited; consequently, the prognosis for these patients is poor. Our aim in the present study was to identify a novel target for antibody therapy against HCC.

Methodology/Principal Findings

We used Western blot and flow cytometric and immunocytochemical analyses to investigate the regulation of FGFR1 expression by interferon-α/β in several human hepatic cancer cell lines. In addition, we tested the efficacy of combined treatment with anti-FGFR1 monoclonal antibody and interferon-α/β in a murine xenograft model of human HCC. We found that interferon-α/β induces expression of FGFR1 in human HCC cell lines, and that an anti-FGFR1 monoclonal antibody (mAb) targeting of the induced FGFR1 can effectively inhibit growth and survival of HCC cells in vitro and in vivo. Moreover, the combination of interferon-α, anti-FGFR1 mAb and peripheral blood mononuclear cells (PBMCs) exerted a significant antitumor effect in vitro.

Conclusions

Our results suggest that the combined use of an anti-FGFR1 antibody and interferon-α/β is a promising approach to the treatment of HCC.     

Involvement of ERK in NMDA receptor-independent cortical neurotoxicity of hydrogen sulfide

Hydrogen sulfide (H₂S), a gasotransmitter, exerts both neurotoxicity and neuroprotection, and targets multiple molecules including NMDA receptors, T-type calcium channels and NO synthase (NOS) that might affect neuronal viability. Here, we determined and characterized effects of NaHS, an H₂S donor, on cell viability in the primary cultures of mouse fetal cortical neurons. NaHS caused neuronal death, as assessed by LDH release and trypan blue staining, but did not significantly reduce the glutamate toxicity. The neurotoxicity of NaHS was resistant to inhibitors of NMDA receptors, T-type calcium channels and NOS, and was blocked by inhibitors of MEK, but not JNK, p38 MAP kinase, PKC and Src. NaHS caused prompt phosphorylation of ERK and upregulation of Bad, followed by translocation of Bax to mitochondria and release of mitochondrial cytochrome c, leading to the nuclear condensation/fragmentation. These effects of NaHS were suppressed by the MEK inhibitor. Our data suggest that the NMDA receptor-independent neurotoxicity of H₂S involves activation of the MEK/ERK pathway and some apoptotic mechanisms.     

Reticulamoeba Is a Long-Branched Granofilosean (Cercozoa) That Is Missing from Sequence Databases

We sequenced the 18S ribosomal RNA gene of seven isolates of the enigmatic marine amoeboflagellate Reticulamoeba Grell, which resolved into four genetically distinct Reticulamoeba lineages, two of which correspond to R. gemmipara Grell and R. minor Grell, another with a relatively large cell body forming lacunae, and another that has similarities to both R. minor and R. gemmipara but with a greater propensity to form cell clusters. These lineages together form a long-branched clade that branches within the cercozoan class Granofilosea (phylum Cercozoa), showing phylogenetic affinities with the genus Mesofila. The basic morphology of Reticulamoeba is a roundish or ovoid cell with a more or less irregular outline. Long and branched reticulopodia radiate from the cell. The reticulopodia bear granules that are bidirectionally motile. There is also a biflagellate dispersal stage. Reticulamoeba is frequently observed in coastal marine environmental samples. PCR primers specific to the Reticulamoeba clade confirm that it is a frequent member of benthic marine microbial communities, and is also found in brackish water sediments and freshwater biofilm. However, so far it has not been found in large molecular datasets such as the nucleotide database in NCBI GenBank, metagenomic datasets in Camera, and the marine microbial eukaryote sampling and sequencing consortium BioMarKs, although closely related lineages can be found in some of these datasets using a highly targeted approach. Therefore, although such datasets are very powerful tools in microbial ecology, they may, for several methodological reasons, fail to detect ecologically and evolutionary key lineages.     

Control of ommochrome synthesis by both juvenile hormone and melanization hormone in the cabbage armyworm,Mamestra brassicae

Summary Pigmentation of last instar larvae of the cabbage armyworm,Mamestra brassicae is of two types: melanin in the cuticle and ommochrome in the epidermis. The latter was found to be primarily xanthommatin. When allatectomy was performed 8 h before head capsule slippage (HCS) in the last larval molt, later ommochrome synthesis was inhibited. Application of juvenile hormone (JH) up to 12 h after HCS (9 h before ecdysis) (activity: methopreneJH I>JH II>JH III) restored ommochrome synthesis. After that time it has less and less effect.Removal of the suboesophageal ganglion from the larvae 8 h before HCS prevented both later ommochrome synthesis and melanization. Melanization of isolated abdomens was restored by implantation of 3 suboesophageal ganglia or injection of melanization and reddish coloration hormone (MRCH) 18 h after HCS. Restoration of ommochrome synthesis required exogenous JH in addition to melanization hormone from suboesophageal ganglion or MRCH. Therefore, melanization appears to be critical for the later onset of ommochrome synthesis even in a larva which has been exposed to JH during the critical period.Abbreviations CC·CA corpora cardiaca-corpora allata complex - JH juvenile hormone - MRCH melanization and reddish coloration hormone - HCS head capsule slippage