Expression of an extracellular ribonuclease gene increases resistance to <Emphasis Type="Italic">Cucumber mosaic virus</Emphasis> in tobacco

Background

The apoplast plays an important role in plant defense against pathogens. Some extracellular PR-4 proteins possess ribonuclease activity and may directly inhibit the growth of pathogenic fungi. It is likely that extracellular RNases can also protect plants against some viruses with RNA genomes. However, many plant RNases are multifunctional and the direct link between their ribonucleolytic activity and antiviral defense still needs to be clarified. In this study, we evaluated the resistance of Nicotiana tabacum plants expressing a non-plant single-strand-specific extracellular RNase against Cucumber mosaic virus.

Results

Severe mosaic symptoms and shrinkage were observed in the control non-transgenic plants 10 days after inoculation with Cucumber mosaic virus (CMV), whereas such disease symptoms were suppressed in the transgenic plants expressing the RNase gene. In a Western blot analysis, viral proliferation was observed in the uninoculated upper leaves of control plants, whereas virus levels were very low in those of transgenic plants. These results suggest that resistance against CMV was increased by the expression of the heterologous RNase gene.

Conclusion

We have previously shown that tobacco plants expressing heterologous RNases are characterized by high resistance to Tobacco mosaic virus. In this study, we demonstrated that elevated levels of extracellular RNase activity resulted in increased resistance to a virus with a different genome organization and life cycle. Thus, we conclude that the pathogen-induced expression of plant apoplastic RNases may increase non-specific resistance against viruses with RNA genomes.

     

Temperature dependence on the pesticide sampling rate of polar organic chemical integrative samplers (POCIS)

Laboratory experiments were performed to determine the sampling rates of pesticides for the polar organic chemical integrative samplers (POCIS) used in Japan. The concentrations of pesticides in aquatic environments were estimated from the accumulated amounts of pesticide on POCIS, and the effect of water temperature on the pesticide sampling rates was evaluated. The sampling rates of 48 pesticides at 18, 24, and 30 °C were obtained, and this study confirmed that increasing trend of sampling rates was resulted with increasing water temperature for many pesticides.     

(Pro)renin receptor accelerates development of sarcopenia via activation of Wnt/YAP signaling axis

To extend life expectancy and ensure healthy aging, it is crucial to prevent and minimize age‐induced skeletal muscle atrophy, also known as sarcopenia. However, the disease's molecular mechanism remains unclear. The age‐related Wnt/β‐catenin signaling pathway has been recently shown to be activated by the (pro)renin receptor ((P)RR). We report here that (P)RR expression was increased in the atrophied skeletal muscles of aged mice and humans. Therefore, we developed a gain‐of‐function model of age‐related sarcopenia via transgenic expression of (P)RR under control of the CAG promoter. Consistent with our hypothesis, (P)RR‐Tg mice died early and exhibited muscle atrophy with histological features of sarcopenia. Moreover, Wnt/β‐catenin signaling was activated and the regenerative capacity of muscle progenitor cells after cardiotoxin injury was impaired due to cell fusion failure in (P)RR‐Tg mice. In vitro forced expression of (P)RR protein in C2C12 myoblast cells suppressed myotube formation by activating Wnt/β‐catenin signaling. Administration of Dickkopf‐related protein 1, an inhibitor of Wnt/β‐catenin signaling, and anti‐(P)RR neutralizing antibody, which inhibits binding of (P)RR to the Wnt receptor, significantly improved sarcopenia in (P)RR‐Tg mice. Furthermore, the use of anti‐(P)RR neutralizing antibodies significantly improved the regenerative ability of skeletal muscle in aged mice. Finally, we show that Yes‐associated protein (YAP) signaling, which is coordinately regulated by Wnt/β‐catenin, contributed to the development of (P)RR‐induced sarcopenia. The present study demonstrates the use of (P)RR‐Tg mice as a novel sarcopenia model, and shows that (P)RR‐Wnt‐YAP signaling plays a pivotal role in the pathogenesis of this disease.     

Structure-activity relationship studies of Bz amide-containing α-GalCer derivatives as natural killer T cell modulators

Abstract

CD1d is a non-polymorphic antigen-presenting glycoprotein that recognizes glycolipids as ligands. Ligands bind to the hydrophobic grooves of CD1d, and the resulting ligand-CD1d complexes activate natural killer T (NKT) cells by means of T cell receptor recognition, leading to the secretion of various cytokines. However, details of the ligand recognition mechanism of a large hydrophobic ligand binding pocket and the relationship between cytokine induction and ligand structure are unclear. We report the synthesis of α-GalCer derivatives containing a Bz amide group having various substituting groups in the ceramide moiety, and the analysis of the structure-activity relationships. The assays reveal that the Bz amide-containing CD1d ligands function as NKT cell modulators displaying Th2 cytokine biasing responses. Furthermore, molecular dynamics simulation studies suggest that the phenyl groups can interact with the aromatic amino acid residues in the lipid binding pocket of CD1d.     

Sclerochronological study of the gigantic inoceramids Sphenoceramus schmidti and S. sachalinensis from Hokkaido,northern Japan

Here, we present the first sclerochronological investigation of shells of the gigantic inoceramids Sphenoceramus schmidti and S. sachalinensis from the middle Campanian cold seep carbonate‐bearing strata of the Yezo Basin in Hokkaido (northern Japan). Stable carbon (δ¹³C) and oxygen (δ¹⁸O) isotope values were measured in the aragonitic and calcitic shell layers of both species and compared to those of other co‐occurring benthic (mainly bivalves and gastropods) and demersal molluscs (ammonites). Sedimentological and stable isotope data suggest that these bivalves lived near cold seeps and were exposed to high H₂S level in the seawater. The inoceramid shells exhibited higher δ¹³C and lower δ¹⁸O values than the coeval non‐cold seep molluscs. We ascribed the anomalous isotopic pattern to a combination of vital and environmental effects determined by the hosting of chemosymbionts and the exposure to warm interstitial waters. Inoceramid δ¹³C minima coincided with growth lines and likely reflect changes in nutrient supply by the chemosymbionts. Absolute temperatures estimated from δ¹⁸O values of Sphenoceramus schmidti and S. sachalinensis were, on average, ca. 4–5°C warmer than those reconstructed for the non‐seepage environment (19.3 ± 0.7°C). Short‐term δ¹⁸O fluctuations of the inoceramid material indicate local temperature ranges of up to 5.2°C, that is four times larger than those reconstructed from the benthic and demersal fauna (1.3°C). In general, our data suggest that the stable carbon and oxygen isotope values of the studied Sphenoceramus spp. were strongly affected by short‐term fluctuations in seepage activity and do not reflect seasonal fluctuations.     

The phenology of cherry blossom (<Emphasis Type="Italic">Prunus yedoensis</Emphasis> “Somei-yoshino”) and the geographic features contributing to its flowering

We investigated relationships between the flowering phenology of Prunus yedoensis "Somei-yoshino" (cherry blossom) and the local temperatures in Japan. Our observations were carried out across the Okayama Plain, which included Okayama City (about 700,000 inhabitants), from the winter of 2008 to the spring of 2009. Local air temperature (AT) and the globe temperature (GT) were recorded at the tree height. The flowering dates (FDs) of P. yedoensis were earliest in the central commercial area (located at the center of the plain), followed by the north residential area (further inland), and finally the south residential area (seaward). The recorded FDs were related to the period-averaged daily maximum/minimum AT and GT, and the phenologically effective AT and GT defined in this study. Of these parameters, the phenologically effective GTs correlated most with the FDs. Since the GT is determined by AT, solar and infrared radiations, and wind speed, our previous result suggests that a combination of these three components surrounding the tree is more important for budding and flowering than is AT alone. The supposition is supported by the flowering of P. yedoensis being the latest at the coastal region of the Okayama Plain where the AT were higher than at the inland region, excluding the urban area; it is probably caused by stronger winds there than at the other sites.     

Subtractive screening of genes involved in cellular senescence

We attempted to identify the genes involved in cellularsenescence, telomere maintenance and telomerase regulationthrough subtractive screening of cDNA libraries prepared froma human lung adenocarcinoma cell line A549 and its sublinesnamed A5DC7, CK and AST-9. Cell phenotypes of A5DC7, CK andAST-9 are normal cell-like, cancer cell-like and intermediate,respectively. These cell lines have different phenotypes interms of telomerase activity and telomere maintenance, andthus are thought to be useful for identifying the genesinvolved in cellular senescence and telomerase regulation. In this study, we identified 86 independent cDNA clones bysubtractive screening. Among these cDNA clones, subtractingA5DC7 cDNAs from A549 cDNAs and CK cDNAs gave 7 and 3 cDNAclones which highly and specifically expressed in tester celllines. Genes corresponding to these 10 cDNA clones mightparticipate in maintaining cancer-cell phenotypes. As aresult of database searching, each four of A549 specific cDNAclones are found to correspond to known cDNAs. Each two ofA549 specific and two of CK specific cDNA clones have highhomology to independent ESTs. Sequences having homology toeach one of A549 specific and one of CK specific cDNA cloneshave not been deposited in the Genbank database, indicatingthat these two cDNA clones are part of novel genes. Weanticipate that their involvement in telomerase regulationand/or senescence program can be clarified by functionalanalysis using each full-length cDNA.     

Effects of Hypoxia on the Activity of the Dopaminergic Neuron System in the Rat Striatum as Studied by In Vivo Brain Microdialysis

The purpose of the present study is to clarify the effects of hypoxia on the activity of the dopaminergic neurons in the brain and its mechanism of action. For this purpose, the effects of hypoxia on the extracellular levels of 3,4-dihy-droxyphenylethylamine (dopamine) were examined in the rat Striatum using in vivo brain microdialysis in the presence or absence of pretreatment with either tetrodotoxin (a blocker of voltage-dependent sodium channels) or nomifensine (a blocker of dopamine reuptake). Exposure to various degrees of hypoxia (15, 10, and 8% O₂ in N₂) increased dopamine levels in striatal dialysates to 200, 400, and 1,100%, respectively, of the control value. On reoxygenation, dopamine levels in the dialysates rapidly returned to the control level. Reexposure to hypoxia increased the dopamine levels to the same extent as during the first exposure. After addition of tetrodotoxin (40 mUM) to the perfusion fluid or pretreatment with nomifensine (100 mg/kg, i.p.), exposure to hypoxia no longer increased the dopamine levels. These results suggest that although hypoxia induces an increase in the extracellular dopamine levels (hence, an apparent increase in the activity of the dopaminergic neurons), this increase is not the result of an increase in dopamine release itself, but rather the result of inhibition of the dopamine reuptake mechanism.