Impacts of land-use history on the diversity of a riparian forest landscape in warm-temperate Kyushu,southern Japan

We examined the impacts of land-use history on the species composition and diversity of a warm-temperate riparian forest landscape in Kyushu, southern Japan, focusing on the relationship between evergreen oaks and deciduous trees in natural and seminatural forests. The species composition of 59 plots was classified into four types (A to D). Type A, which showed a significant bias towards sites not subject to nonforest land use since 1947, had high species diversity consisting of (1) many lucidophyllous components of the region, including the rare indigenous oak Quercus hondae, and (2) summergreen tree species of varying dominance and number representing unique or locally rare elements of the riparian landscape in this warm-temperate region. Type B was dominated by a common species of oak, Q. glauca, and displayed less clear distribution bias with land-use history. In contrast to types A and B, types C and D, which were characterized by high dominance of deciduous trees, had negative bias away from sites that had been under forest land use in 1947. Presumably, intensive anthropogenic disturbances associated with nonforest land uses had expanded the habitats for deciduous trees. This phenomenon was represented by the establishment of forests (type D) dominated by Ulmus davidiana var. japonica (UDJ) after it had been released from the suppression of evergreen forest trees during a period of nonforest land use that prevents the successful recovery of evergreen trees. From these results we conclude that the impacts of land-use history on the diversity of warm-temperate riparian forest landscape are multiphased: a period of nonforest land use has a strong negative impact on lucidophyllous forest trees represented by the rare indigenous oak Q. hondae; release from the suppressive effects of the lucidophyllous species then encourages establishment of locally rare deciduous tree flora represented by UDJ, which continue to persist for decades after abandonment of nonforest land use.     

Detection of Pairwise Residue Proximity by Covariation Analysis for 3D-Structure Prediction of G-Protein-Coupled Receptors

G-protein-coupled receptor (GPCR) is one of the most important targets for medicines. Homology modeling based on the crystal structure of bovine rhodopsin is currently the most frequently used method for GPCR targeted drug design. Information about residue-residue contacts and the structural specificity in the subfamily is essential for constructing more precise 3D structures, to distinguish the structural differences between the template and targets. In this study, we adopted the covariation analysis to extract information about residue-residue interactions from the amino acid sequence. In the opsin family, a large number of adjacent covarying residue pairs were detected. The detected residue pairs have a strong tendency to gather in some regions important for the structure and function. These results suggest that the covariation analysis is practically utilized to detect adjacent residue pairs and also to apply for predicting functional sites. Analyses of other GPCR subfamilies, olfactory receptor and chemokine receptor families, demonstrated that some adjacent covarying residue pairs were common. Thus, the covariation analysis has possibilities in the substantial improvement of the 3D-structure modeling of GPCRs and in the detection of functional sites such as the ligand-binding sites.     

MSLN Gene Silencing Has an Anti-Malignant Effect on Cell Lines Overexpressing Mesothelin Deriving from Malignant Pleural Mesothelioma

Genes involved in the carcinogenetic mechanisms underlying malignant pleural mesothelioma (MPM) are still poorly characterized. So far, mesothelin (MSLN) has aroused the most interest. It encodes for a membrane glycoprotein, frequently over-expressed in various malignancies such as MPM, and ovarian and pancreatic cancers. It has been proposed as a diagnostic and immunotherapeutic target with promising results. However, an alternative therapeutic approach seems to rise, whereby synthetic molecules, such as antisense oligonucleotides, could be used to inhibit MSLN activity. To date, such a gene-level inhibition has been attempted in two studies only, both on pancreatic and ovarian carcinoma cell lines, with the use of silencing RNA approaches. With regard to MPM, only one cell line (H2373) has been employed to study the effects of MSLN depletion. Indeed, the knowledge on the role of MSLN in MPM needs expanding. Accordingly, we investigated the expression of MSLN in a panel of three MPM cell lines, i.e. NCI-H28, Mero-14, and IstMes2; one non-MPM cell line was used as reference (Met5A). MSLN knock-down experiments on MSLN-overexpressing cells were also performed through silencing RNA (siRNA) to verify whether previous findings could be generalized to a different set of cell cultures. In agreement with previous studies, transient MSLN-silencing caused decreased proliferation rate and reduced invasive capacity and sphere formation in MSLN-overexpressing Mero-14 cells. Moreover, MSLN-siRNA combined with cisplatin, triggered a marked increase in apoptosis and a decrease in proliferation as compared to cells treated with each agent alone, thereby suggesting a sensitizing effect of siRNA towards cisplatin. In summary, our findings confirm that MSLN should be considered a key molecular target for novel gene-based targeted therapies of cancer.     

Flavonoids induce germination of basidiospores of the ectomycorrhizal fungus <Emphasis Type="Italic">Suillus bovinus</Emphasis>

Under laboratory conditions, spores of ectomycorrhizal fungi usually germinate very poorly or not at all. In a previous study, we showed that spores of the ectomycorrhizal fungus Suillus bovinus germinated through the combination of activated charcoal treatment of media and co-culture with seedlings of Pinus densiflora, which suggested that some substances contained in root exudates induced the germination. Among the compounds reported from root exudates, flavonoids have been elucidated to play various and substantial roles in plant–microbe interactions; we therefore investigated the effects of flavonoids on basidiospore germination of S. bovinus by the diffusion gradient assay on water agar plates pretreated with charcoal powder. Seven out of the 11 flavonoids tested, hesperidin, morin, rutin, quercitrin, naringenin, genistein, and chrysin, had greater effects than controls, whereas flavone, biochanin A, luteolin, and quercetin showed no positive effects. The effective concentration presumably corresponded to several micromolar levels, which was equivalent to those effective for pollen development, nod gene induction, and spore germination of F. solani f. sp. pisi and AM fungi. The results suggest that flavonoids play a role as signaling molecules in symbiotic relationships between woody plants and ectomycorrhizal fungi.     

Immunostimulatory activity of major membrane protein-II from Mycobacterium leprae

We examined the antigenicity of an immunomodulatory protein, major membrane protein (MMP)-II, from Mycobacterium leprae, since host defense against M. leprae largely depends on adaptive immunity. Both unprimed and memory T cells from healthy individuals were stimulated by autologous MMP-II-pulsed monocyte-derived dendritic cells (DCs) to produce IFN-gamma. The DC-mediated IFN-gamma production was dependent on the expression of MHC, CD86, and MMP-II antigens. Memory T cells from paucibacillary (PB) leprosy more extensively responded to MMP-II-pulsed DCs than T cells from healthy individuals, while comparable IFN-gamma was produced by unprimed T cells. Memory T cells from multibacillary leprosy, which are normally believed to be anergic, were activated similarly to those from healthy individuals by MMP-II-pulsed DCs. These results suggest that memory T cells from PB leprosy are primed with MMP-II prior to the manifestation of the disease, and MMP-II is highly antigenic in terms of activation of adaptive immunity.     

Extramembrane central pore of multidrug exporter AcrB in Escherichia coli plays an important role in drug transport

We previously reported the crystal structure of the major multidrug exporter AcrB in Escherichia coli (Murakami, S., Nakashima, R., Yamashita, E., and Yamaguchi, A. (2002) Nature 419, 587-593). The extramembrane headpiece of the AcrB trimer contains a central pore composed of three alpha-helices. Each pore helix belongs to a different monomer. In this study, we constructed cysteine-scanning mutants as to the residues comprising the pore helix. Of the 21 mutants (D99C to P119C), 5 (D101C, V105C, N109C, Q112C, and P116C) showed significantly reduced drug resistance and drug-exporting activity. These residues are localized on one side of the pore helix, i.e. on the wall of the pore. These observations strongly indicate the important role of this pore in the drug transport process. A N-ethylmaleimide binding experiment revealed that the pore is in the closed state, and the thickness of the permeability barrier in the middle of the pore corresponds to 2.5 alpha-helical turns. Two mutants (V105C and Q112C), which showed the greatest loss of activity of all of the pore mutants, were detected in the form of disulfide cross-linking dimers under normal conditions, suggesting that a conformational change of the pore is indispensable during the transport process.     

Polycomb group protein Ezh1 represses Nodal and maintains the left-right axis

The left-right (LR) axis is essential for the proper function of internal organs. In mammals and fish, left-sided Nodal expression governs LR patterning. Here, we show that the Polycomb group protein Ezh1, which is highly conserved from fish to human, participates in LR patterning. Knockdown of olezh1, a medaka homologue of Ezh1, led to LR reversal of internal organs. It was shown that OLEZH1 acts in silencing the expression of Spaw (a medaka homolog of Nodal) via a previously unknown pathway. Furthermore, coimmunoprecipitation showed physical interaction of Ezh1 with FoxH1, a Nodal regulator. This represents a novel mechanism for LR patterning and implies that Ezh1 has developmental importance.     

First report of Mycopappus alni in Japan: species identification of the pathogenic fungus of a frosty mildew disease in Crataegus chlorosarca

A leaf disease similar to frosty mildew disease caused by Mycopappus species was detected on the leaves of Crataegus chlorosarca in Tomakomai, Hokkaido Prefecture. From morphological observations and gene analyses of rDNA-ITS, the fungus was identified as M. alni, which causes leaf blight disease on Alnus spp., Betula spp., and a Pyrus sp. in North America and Turkey. This is the first report of M. alni in Japan and Crataegus as its new host genus.