Studies on the Mode of Action of Organophosphorus Compounds

For the purpose of distinguihsing sumithion from methylparathion in the mammalian metabolism, phosphorus³² labeled compounds were administered to Guinea pig and white rat. Both compounds were found to be absorbed readily, and phosphorus containing metabolites excreted chiefly into urine. By chromatographic separation and identification of the metabolites, the decomposition of sumithion was observed to proceed presumably more easily than methylparathion. From these results lower toxicity of the former toward mammals than the latter was discussed. In addition, compounds remaining in rice plant and German cockroach were also analysed.     

C-Mannosylated peptides derived from the thrombospondin type 1 repeat enhance lipopolysaccharide-induced signaling in macrophage-like RAW264.7 cells

C-Mannosylation is a unique type of glycosylation occurring at the first Trp (W) in the WXXW motif, which is found in the thrombospondin type 1 repeat (TSR) of proteins. However, the biological function of C-mannosylation is not fully understood. In this study, we investigated the effect of C-mannosylated TSR-derived peptides on lipopolysaccharide (LPS)-induced signaling in macrophage-like RAW264.7 cells. The cells were stimulated with LPS in the presence or absence of chemically synthesized peptides with or without C-mannose (e.g., (C-Man)-Trp-Ser-Pro-Trp [C-Man-WSPW], C-Man-W, WSPW, etc.), then the effects of the peptides on cellular viability and signaling were examined. We found a cytotoxic effect in the cells treated with LPS and C-Man-WSPW, but not in the cells solely treated with LPS or C-Man-WSPW. We also found that production of tumor necrosis factor-alpha (TNF-alpha) was upregulated more in response to LPS plus C-Man-WSPW, than in response to LPS plus WSPW or LPS alone. Among the LPS-induced signaling pathways that induce production of TNF-alpha, the activation of c-Jun N-terminal kinase (JNK) was greatly enhanced by LPS and C-Man-WSPW, and the production of TNF-alpha was suppressed by an inhibitor for JNK. Together, these results demonstrate a novel function of the C-mannosylated TSR-derived peptide motif, to promote LPS-induced JNK signaling, and this leads to an enhancement of cytotoxicity via the upregulation of TNF-alpha production.     

The potential benefits of nicaraven to protect against radiation-induced injury in hematopoietic stem/progenitor cells with relative low dose exposures

Nicaraven, a hydroxyl radical-specific scavenger has been demonstrated to attenuate radiation injury in hematopoietic stem cells with 5 Gy γ-ray exposures. We explored the effect and related mechanisms of nicaraven for protecting radiation injury induced by sequential exposures to a relatively lower dose γ-ray. C57BL/6 mice were given nicaraven or placebo within 30 min before exposure to 50 mGy γ-ray daily for 30 days in sequences (cumulative dose of 1.5 Gy). Mice were victimized 24 h after the last radiation exposure, and the number, function and oxidative stress of hematopoietic stem cells were quantitatively estimated. We also compared the gene expression in these purified stem cells from mice received nicaraven and placebo treatment. Nicaraven increased the number of c-kit⁺ stem/progenitor cells in bone marrow and peripheral blood, with a recovery rate around 60–90% of age-matched non-irradiated healthy mice. The potency of colony forming from hematopoietic stem/progenitor cells as indicator of function was completely protected with nicaraven treatment. Furthermore, nicaraven treatment changed the expression of many genes associated to DNA repair, inflammatory response, and immunomodulation in c-kit⁺ stem/progenitor cells. Nicaraven effectively protected against damages of hematopoietic stem/progenitor cells induced by sequential exposures to a relatively low dose radiation, via complex mechanisms.     

Enhanced Nox1 expression and oxidative stress resistance in c-kit-positive hematopoietic stem/progenitor cells

Although stem cells are generally thought to be resistant to oxidative stress, the fact and in detail molecular mechanism are still to be clearly identified. We herein tried to understand the overall characterization of redox regulatory signaling in hematopoietic stem cells. We purified c-kit-positive hematopoietic stem/progenitor cells from the bone marrow of healthy mice, and then evaluated their redox regulatory property. Compared to the c-kit-negative matured mononuclear cells, c-kit-positive stem/progenitor cells showed lower basic levels of intracellular reactive oxygen species, faster clearance of the accumulated intracellular reactive oxygen species, and higher resistant to oxidative stress. An overall view on the gene expression profile associated with redox regulation showed to be widely differed between cell types. We confirmed that the c-kit-positive stem/progenitor cells expressed significantly higher of Nox1 and catalase, but less of lactoperoxidase than these matured mononuclear cells. Our data suggests that stem cells keep specific redox regulatory property for defensing against oxidative stress.     

Ralstonia solanacearum type III secretion system effector Rip36 induces a hypersensitive response in the nonhost wild eggplant Solanum torvum

Ralstonia solanacearum is a Gram‐negative soil‐borne bacterium that causes bacterial wilt disease in more than 200 plant species, including economically important Solanaceae species. In R. solanacearum, the hypersensitive response and pathogenicity (Hrp) type III secretion system is required for both the ability to induce the hypersensitive response (HR) in nonhost plants and pathogenicity in host plants. Recently, 72 effector genes, called rip (Ralstonia protein injected into plant cells), have been identified in R. solanacearum RS1000. RS1002, a spontaneous nalixidic acid‐resistant derivative of RS1000, induced strong HR in the nonhost wild eggplant Solanum torvum in an Hrp‐dependent manner. An Agrobacterium‐mediated transient expression system revealed that Rip36, a putative Zn‐dependent protease effector of R. solanacearum, induced HR in S. torvum. A mutation in the putative Zn‐binding motif (E149A) completely abolished the ability to induce HR. In agreement with this result, the RS1002‐derived Δrip36 and rip36E149A mutants lost the ability to induce HR in S. torvum. An E149A mutation had no effect on the translocation of Rip36 into plant cells. These results indicate that Rip36 is an avirulent factor that induces HR in S. torvum and that a putative Zn‐dependent protease motif is essential for this activity.     

Two members of the IgLON family are expressed in a restricted region of the developing chick brain and neural crest

The precise expression patterns of two IgLON genes, CEPU-1 and limbic system-associated membrane protein (LAMP), were studied during early embryogenesis. It was found that expression of both was localized to restricted regions of the brain and neural crest. In the developing neural tube, CEPU-1 was expressed in the isthmus and a restricted region of the hindbrain, whereas LAMP was expressed in the anterior midbrain. Most neural crest cells expressed LAMP, whereas CEPU-1 expression was limited to crest cells derived from the hindbrain. These results suggest that members of the IgLON family have important roles during embryogenesis, particularly in brain formation and differentiation.     

Flagellin glycosylation island in Pseudomonas syringae pv. glycinea and its role in host specificity

The deduced amino acid sequences of the flagellins of Pseudomonas syringae pv. tabaci and P. syringae pv. glycinea are identical; however, their abilities to induce a hypersensitive reaction are clearly different. The reason for the difference seems to depend on the posttranslational modification of the flagellins. To investigate the role of this posttranslational modification in the interactions between plants and bacterial pathogens, we isolated genes that are potentially involved in the posttranslational modification of flagellin in P. syringae pv. glycinea (glycosylation island); then defective mutants with mutations in these genes were generated. There are three open reading frames in the glycosylation island, designated orf1, orf2, and orf3. orf1 and orf2 encode putative glycosyltransferases, and mutants with defects in these open reading frames, deltaorf1 and deltaorf2, secreted nonglycosylated and slightly glycosylated flagellins, respectively. Inoculation tests performed with these mutants and original nonhost tobacco leaves revealed that deltaorf1 and deltaorf2 could grow on tobacco leaves and caused symptom-like changes. In contrast, these mutants failed to cause symptoms on original host soybean leaves. These data indicate that putative glycosyltransferases encoded in the flagellin glycosylation island are strongly involved in recognition by plants and could be the specific determinants of compatibility between phytopathogenic bacteria and plant species.     

On the Mechanism of Radiation Enhancement of Lethal Effect of Sodium Chloride on Microorganisms

In connection with the preceding paper concerning the enhancement of radiation lethal effect by the NaCl treatment, the experimental evidences and the further discussion for the possible mechanism of the enhancement by the post-NaCl treatment are presented.

In order to elucidate the radiation damages of cell constituents and its enhancement by NaCl, the infrared analysis and isotopic experiments were carried out with cells of S. cerevisiae, Z. soya and E. coli.

Consequently, it has been concluded that the possible mechanism may involve the processes of increase in cell permeability by irradiation and of the denaturation of the critical cell constituents by increasing penetration of NaCl and thus the sensitivity of irradiated cells to NaCl may increase. The NaCl sensitive sites are considered not to be restricted to the biological targets for radiation inactivation by the supports of the survival curve analysis based on the target theory and of the isotopic experiment.     

Efficient gene targeting by homologous recombination in rice

Modification of genes through homologous recombination, termed gene targeting, is the most direct method to characterize gene function. In higher plants, however, the method is far from a common practice. Here we describe an efficient and reproducible procedure with a strong positive/negative selection for gene targeting in rice, which feeds more than half of the world's population and is an important model plant. About 1% of selected calli and their regenerated fertile plants were heterozygous at the targeted locus, and only one copy of the selective marker used was found at the targeted site in their genomes. The procedure's applicability to other genes will make it feasible to obtain various gene-targeted lines of rice.