Aminooxy‐naphthylpropionic acid and its derivatives are inhibitors of auxin biosynthesis targeting l‐tryptophan aminotransferase: structure–activity relationships

We previously reported l ‐α‐aminooxy‐phenylpropionic acid (AOPP) to be an inhibitor of auxin biosynthesis, but its precise molecular target was not identified. In this study we found that AOPP targets TRYPTOPHAN AMINOTRANSFERASE of ARABIDOPSIS 1 (TAA1). We then synthesized 14 novel compounds derived from AOPP to study the structure–activity relationships of TAA1 inhibitors in vitro. The aminooxy and carboxy groups of the compounds were essential for inhibition of TAA1 in vitro. Docking simulation analysis revealed that the inhibitory activity of the compounds was correlated with their binding energy with TAA1. These active compounds reduced the endogenous indole‐3‐acetic acid (IAA) content upon application to Arabidopsis seedlings. Among the compounds, we selected 2‐(aminooxy)‐3‐(naphthalen‐2‐yl)propanoic acid (KOK1169/AONP) and analyzed its activities in vitro and in vivo. Arabidopsis seedlings treated with KOK1169 showed typical auxin‐deficient phenotypes, which were reversed by exogenous IAA. In vitro and in vivo experiments indicated that KOK1169 is more specific for TAA1 than other enzymes, such as phenylalanine ammonia‐lyase. We further tested 41 novel compounds with aminooxy and carboxy groups to which we added protection groups to increase their calculated hydrophobicity. Most of these compounds decreased the endogenous auxin level to a greater degree than the original compounds, and resulted in a maximum reduction of about 90% in the endogenous IAA level in Arabidopsis seedlings. We conclude that the newly developed compounds constitute a class of inhibitors of TAA1. We designated them ‘pyruvamine’.     

Soil spore banks of ectomycorrhizal fungi in endangered Japanese Douglas-fir forests

Interactions between trees and ectomycorrhizal fungi are critical to the growth and survival of both partners. However, ectomycorrhizal symbiosis has barely been explored in endangered trees, and no information is available regarding soil spore banks of ectomycorrhizal fungi from forests of threatened trees. Here, we evaluated soil spore banks of ectomycorrhizal fungi from endangered Japanese Douglas-fir (Pseudotsuga japonica) forests using bioassay approaches with congeneric P. menziesii and Pinus densiflora seedlings in combination with molecular identification techniques. Rhizopogon togasawariana was predominant in soil propagule banks and was found in all remaining P. japonica forests when assayed with P. menziesii, while no colonization of this fungus was observed on Pinus seedlings. Given the observed specificity of R. togasawariana for P. menziesii and its phylogenetic position within the Pseudotsuga-specific Rhizopogon lineage, its geographical distribution is likely restricted to the remaining Japanese Douglas-fir forests, indicating a high extinction risk for this fungus as well as its endangered host. Spore banks of R. togasawariana remained highly infective after preservation for 1 year or heat treatment at 70 °C, suggesting an ecological strategy of establishing ectomycorrhizal associations on regenerating Japanese Douglas-fir seedlings after disturbance, as observed in other Rhizopogon–Pinaceae combinations. Therefore, the regeneration of Japanese Douglas-fir seedlings may depend largely on the soil spore banks dominated by R. togasawariana, which has co-evolved with the Japanese Douglas-fir for over 30 million years. More attention must be paid to underground ectomycorrhizal fungi for the conservation of endangered tree species, especially in the era of human-induced mass extinction.     

Environmental factors determining the distribution of highland plants at low-altitude algific talus sites

Algific talus is a micro-scale habitat type where highland plants (subalpine and alpine species) are found, disjunct from their typical range, in lowland forests. On algific talus, cold airflows from the interstices between talus fragments create a local microclimate colder than surrounding forests. Despite of the widely-known occurrence of unique vegetation on algific talus, critical environmental factors determining the distribution of highland species in this habitat type are unclear. In order to reveal the environmental factors enabling highland species to inhabit algific talus, we investigated the vegetation and environments of 26 algific talus sites and four reference (non-algific talus) sites in Hokkaido, northern Japan. Several algific talus sites were dominated by highland species, while some algific talus sites and all non-algific talus sites were dominated by lowland species. Community analysis based on detrended correspondence analysis (DCA) and canonical corresponding analysis (CCA) revealed that the algific talus sites dominated by highland species had lower ground temperature, more acidic soil, larger canopy openness, and less diverse vegetation than the sites dominated by lowland species. Highland plants might be maintained under conditions stressful for lowland plants, resulting in less competitive situation. Generalized linear models (GLM), used to evaluate the response of individual highland species to environmental factors, revealed that preferable environmental conditions for highland plants are highly species specific. These results indicate that the maintenance of diverse environments is crucial for the conservation of the unique vegetation and local populations of highland species in algific talus areas.     

Insertion hot spot for horizontally acquired DNA within a bidirectional small-RNA locus in Salmonella enterica

In Escherichia coli and Salmonella enterica, RyeA and RyeB RNAs are encoded on opposite DNA strands at the same locus. We present evidence indicating that the last 23 bp of the ryeB gene, corresponding to an internal portion of the ryeA gene, served repeatedly as the integration site for exogenous DNA during Salmonella evolution and still act as an attachment site for present-day bacteriophages. Interestingly, ryeA sequence and expression are modified upon lysogenization.     

Phylogenetic diversity of non-nodulating Rhizobium associated with pine ectomycorrhizae

Most Rhizobium species described are symbionts that form nodules on legume roots; however, non-nodulating strains of Rhizobium are also widespread in nature. Unfortunately, knowledge of non-nodulating Rhizobium is quite limited compared with nodulating Rhizobium . Here, we studied the phylogenetic diversity of Rhizobium species that inhabit Japanese red pine roots ( Pinus densiflora ). Because fine roots of pine trees are usually colonized by ectomycorrhizal fungi in nature, we mainly used ectomycorrhizal root tips for bacterial isolation. Out of 1195 bacteria isolated from 75 independent root samples from the field and greenhouse experiments, 102 isolates were confirmed to be Rhizobium following partial 16S rRNA gene analysis. Rhizobium species were occasionally dominant in culturable bacterial communities, whereas no Rhizobium species were isolated from the soil itself. Molecular phylogenetic analyses using 16S rRNA, atpD , and recA gene sequences revealed that isolated Rhizobium strains were phylogenetically diverse and that several were distantly related to known Rhizobium species. Considering that a single species of pine is associated with unique and phylogenetically diverse Rhizobium populations, we should pay more attention to non-nodulating strains to better understand the diversity, ecology, and evolution of the genus Rhizobium and plant– Rhizobium associations.     

Proteome approach for identification of schistosomiasis japonica vaccine candidate antigen

Experimental vaccination with radiation-attenuated cercariae (RAC) confers possible practical levels of resistance to challenge infection by humoral and by cellular mechanism. Here, we aimed to identify possible vaccine antigens by using specific IgG antibody from RAC vaccinated miniature pig. Two milligrams of soluble egg antigen (SEA) or schistosomal worm antigen preparation (SWAP) was fractionated using two dimensional liquid chromatography (proteome PF 2D) consisted of high performance chromatofocusing (HPCF) and high resolution reversed phase chromatography (HPRP). Of the 42 HPCF fractions of SEA or SWAP, 26 (61.9%) or 15 (35.7%) showed positive dot blot reaction with RAC vaccinated serum respectively. The dot blot positive fractions were applied to the second HPRP column. One hundred and seven out of 26 x 96 of SEA fractions and 18 out of 15 x 96 SWAP fractions reacted with RAC vaccinated serum. From the positive fractions we chose 17 of SEA and 10 of SWAP that had no reactivity with normal cercariae infected (NCI) sera and had single peak of 214 nm; and automated N-terminal amino acid sequence based on in situ Edman Reaction was conducted. Four sequences were obtained and applied to the homology search in NCBI database. A total of eight candidate genes were listed up and their cDNA clones from schistosomula stage were obtained. Two of the recombinant proteins (AAW27472.1 and AXX25883.1) showed strong reactivity with the RAC vaccinated serum but marginal with NCI serum. This protocol using proteome PF 2D could be applicable in identifying immunoreactive proteins from crude extract for the development of vaccines or for diagnostics.     

Effects of fine sediment accumulation on the redd environment and the survival rate of masu salmon (Oncorhynchus masou) embryos

In recent years, fine sediment, produced by run-off associated with forestry activity and agricultural development that accumulates on riverbeds, has exerted a deleterious influence on lotic ecosystems. This study examined the Oroennukibetsu River, a tributary of the Nukibetsu River, which has been affected by high loads of suspended sediments. Effects of accumulation of fine sediment on the survival rate of masu salmon embryo and also on the redd environment (permeability and intragravel dissolved oxygen concentration) were quantified through a field experiment. Results show that the interchange of DO between intragravel and surface water was not affected directly by permeability or the accumulated fine sediment and that intragravel flow rates can be an important factor controlling embryo survival. A decrease in permeability associated with accumulation of fine sediment lowered the survival rate of embryos by suffocation because the flux of DO that should be supplied to the embryo was severely limited. This situation might be created by the combined effects of an accumulation of fine sediment on the redd and a low DO concentration in the surface water because the DO concentration almost coincided with the intragravel DO.     

Increased Carbohydrate Induced Ghrelin Secretion in Obese vs. Normal‐weight Adolescent Girls

Orexigenic and anorexigenic pathways mediate food intake and may be affected by meal composition. Our objective was to determine whether changes in levels of active ghrelin and peptide YY (PYY) differ in obese vs. normal‐weight adolescent girls following specific macronutrient intake and predict hunger and subsequent food intake. We enrolled 26 subjects: 13 obese and 13 normal‐weight girls, 12–18 years old, matched for maturity (as assessed by bone age) and race. Subjects were assigned a high‐carbohydrate, high‐protein, and high‐fat breakfast in random order. Active ghrelin and PYY were assessed for 4 h after breakfast and 1 h after intake of a standardized lunch. Hunger was assessed using a standardized visual analog scale (VAS). No suppression in active ghrelin levels was noted following macronutrient intake in obese or normal‐weight girls. Contrary to expectations, active ghrelin increased in obese girls following the high‐carbohydrate breakfast, and the percent increase was higher than in controls (P = 0.046). Subsequent food intake at lunch was also higher (P = 0.03). Following the high‐fat breakfast, but not other breakfasts, percent increase in PYY was lower (P = 0.01) and subsequent lunch intake higher (P = 0.005) in obese compared with normal‐weight girls. In obese adolescents, specific intake of high‐carbohydrate and high‐fat breakfasts is associated with greater increases in ghrelin, lesser increases in PYY, and higher intake at a subsequent meal than in controls. Changes in anorexigenic and orexigenic hormones in obese vs. normal‐weight adolescents following high‐carbohydrate and high‐fat meals may influence hunger and satiety signals and subsequent food intake.     

Activation of nucleotide‐binding domain‐like receptor containing protein 3 inflammasome in dendritic cells and macrophages by Streptococcus sanguinis

Streptococcus sanguinis is frequently isolated from the blood of patients with infective endocarditis and contributes to the pathology of this disease through induction of interleukin (IL)‐1β responsible for the development of the disease. However, the mechanism of IL‐1β induction remains unknown. In this study, S. sanguinis activated a murine dendritic cell (DC) to induce IL‐1β and this activity was attenuated by silencing the mRNAs of nucleotide‐binding domain‐like receptor containing protein 3 (NLRP3) and caspase‐1. S. sanguinis induced IL‐1β production in murine bone marrow‐derived macrophage, but this activity was significantly reduced in bone marrow‐derived macrophages from NLRP3‐, apoptosis‐associated speck‐like protein containing a caspase‐recruitment domain‐, and caspase‐1‐deficient mice. DC phagocytosed S. sanguinis cells, followed by the release of adenosine triphosphate (ATP). The ATP‐degradating enzyme attenuated the release of ATP and IL‐1β. The inhibitors for ATP receptor reduced IL‐1β release in DC. These results strongly suggest that S. sanguinis has the activity to induce IL‐1β through the NLRP3 inflammasome in macrophage and DC and interaction of purinergic receptors with ATP released is involved in expression of the activity.