Land plants produce diverse flavonoids for growth, survival, and reproduction. Chalcone synthase is the first committed enzyme of the flavonoid biosynthetic pathway and catalyzes the production of ...2',4,4',6'-tetrahydroxychalcone (THC). However, it also produces other polyketides, including p-coumaroyltriacetic acid lactone (CTAL), because of the derailment of the chalcone-producing pathway. This promiscuity of CHS catalysis adversely affects the efficiency of flavonoid biosynthesis, although it is also believed to have led to the evolution of stilbene synthase and p-coumaroyltriacetic acid synthase. In this study, we establish that chalcone isomerase-like proteins (CHILs), which are encoded by genes that are ubiquitous in land plant genomes, bind to CHS to enhance THC production and decrease CTAL formation, thereby rectifying the promiscuous CHS catalysis. This CHIL function has been confirmed in diverse land plant species, and represents a conserved strategy facilitating the efficient influx of substrates from the phenylpropanoid pathway to the flavonoid pathway.
Near-infrared (NIR) fluorescent imaging is a powerful tool for the non-invasive visualization of the inner structure of living organisms. Recently, NIR fluorescence imaging at 1000-1400 nm (second ...optical window) has been shown to offer better spatial resolution compared with conventional NIR fluorescence imaging at 700-900 nm (first optical window). Here we report lead sulfide (PbS) quantum dots (QDs) and their use for in vivo NIR fluorescence imaging of cerebral venous thrombosis in septic mice. Highly fluorescent PbS QDs with a 1100 nm emission peak (QD1100) were prepared from lead acetate and hexamethyldisilathiane, and the surface of QD1100 was coated with mercaptoundecanoic acid so as to be soluble in water. NIR fluorescence imaging of the cerebral vessels of living mice was performed after intravascular injection (200-300 μL) of QD1100 (3 μM) from a caudal vein. By detecting the NIR fluorescence of QD1100, we achieved non-invasive NIR fluorescence imaging of cerebral blood vessels through the scalp and skull. We also achieved NIR fluorescence imaging of cerebral venous thrombosis in septic mice induced by the administration of lipopolysaccharide (LPS). From the NIR fluorescence imaging, we found that the number of thrombi in septic mice was significantly increased by the administration of LPS. The formation of thrombi in cerebral blood vessels in septic mice was confirmed by enzyme-linked immunosorbent assay (ELISA). We also found that the number of thrombi significantly decreased after the administration of heparin, an inhibitor of blood coagulation. These results show that NIR fluorescence imaging with QD1100 is useful for the evaluation of the pathological state of cerebral blood vessels in septic mice.
An edible gall is formed between the third and fourth nodes beneath the apical meristem near the base of
Zizania latifolia
shoots. This gall is harbored by and interacts with the smut fungus
Ustilago ...esculenta
. The gall is also a valuable vegetable called “white bamboo,”
jiaobai
or
gausun
in China and
makomotake
in Japan. Five samples of the galls harvested at different stages of swelling were used to isolate microorganisms by culturing. Isolated fungal and bacterial colonies were identified by DNA sequencing and matrix-assisted laser desorption/ionization–time-of-flight mass spectrometry, respectively. Several strains of
U. esculenta
as well as 6 other species of fungi and 10 species of bacteria were isolated. The microbiome was also evaluated by simple and outlined DNA profiling with automated rRNA intergenic spacer analysis (ARISA), and the amount of DNA of
U. esculenta
was determined by qPCR. At least 16 species of fungi and 40 species of bacteria were confirmed by ARISA of the overall sample. Interestingly, the greatest bacterial diversity, i.e., 18 species, was observed in the most mature sample, whereas the fungal diversity observed in this sample, i.e., 4 species, was rather poor. Based on qPCR,
U. esculenta
occurred in samples from all stages; however, the abundance of
U. esculenta
exhibited unique U-shaped relationships with growth. These results may explain why the interaction between
U. esculenta
and
Z. latifolia
also influences the unique microbial diversity observed throughout the growth stages of the swollen shoot, although the limited sample size does not allow conclusive findings.
Larvae of many odonate species commonly appear within the same freshwater ecosystem. However, it remains unknown whether there is a dietary separation among the larvae of different species. In this ...study, we collected different species of odonate larvae in different seasons at various sites on the floating mat of Mizorogaike Pond in Kyoto, Japan, and examined the abundance and gut contents of the dominant species. Our analyses showed that habitat and seasonal abundance of the odonate larvae did not significantly overlap between most pairs of the dominant species. Moreover, the degree of dietary overlap was not related to larval biomass in microhabitats. These results suggest that, although larval food habits were similar among species, the existence of various vegetation types on the floating mat allowed many odonate species to inhabit Mizorogaike Pond.
Because littoral cladocerans are common prey for odonate larvae, they are ideal organisms to examine prey and predator interactions in aquatic habitats. Although death‐feigning behaviour observed in ...littoral cladocerans is viewed as a trait to reduce predation risk, no study has examined the effectiveness of death‐feigning at reducing mortality. We studied the effectiveness of death‐feigning behaviour by two littoral cladocerans, Chydorus sphaericus and Oxyurella tenuicaudis, against the odonate larvae of Sympetrum frequens.
We first examined how the odonate larvae detected the prey and found that the larvae consumed active cladocerans, even under dark conditions, but they did not consume anaesthetised cladocerans, indicating that the larvae detect the prey using mechanical cues such as vibrational currents created by the prey individuals.
We then observed behavioural events of odonate larvae and prey cladocerans using a video‐recording system to count the number of cladocerans that showed death‐feigning behaviours when odonate larvae attacked and those that were not consumed when they showed death‐feigning behaviours.
We observed a total of 1,099 behavioural events in prey–predator interactions. On average, the odonate larvae pursued 63% of the cladocerans encountered, attacked 89% of the cladocerans pursued, and successfully captured only 29% of the cladocerans with the first attack. Among the cladocerans that were not captured, 38% of the individuals continually swam, but 62% ceased to move and exhibited death‐feigning behaviours. The odonate larvae's capture rate for cladocerans exhibiting the death‐feigning behaviours fell significantly short of that for cladocerans that were continually swimming.
The experiments clearly showed that the littoral cladocerans' death‐feigning behaviour effectively decreased predation rate by odonate larvae. The results suggest that the death feigning serves as an adaptive behaviour to reduce mortality risk in littoral cladocerans under predation by such predators that can detect prey using mechanical cues such as those of vibrational currents.
White bamboo is the swollen stem of Zizania latifolia parasitized by the smut fungus Ustilago esculenta. Five samples of galls were harvested at different stages of swelling, along with young ...seedlings of Z. latifolia and isolated colonies of U. esculenta. The inhibition capacity of boiling water or 50% ethanol extracts on NO release by RAW264 cells, which was stimulated with lipopolysaccharide, was evaluated as anti-inflammatory activity, while the NO induction capacity was evaluated as immunostimulatory activity. Total polyphenol (TPP) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging activity were also measured. The anti-inflammatory effect, as well as TPP and DPPH, was comprehensively detected in the ethanol extracts. The immunostimulating activity was observed in the boiled extracts at different levels, depending on the swelling stage, and was especially high in the top (apical) part. These data may indicate that functional components could be dynamically induced by interactions between Z. latifolia and U. esculenta.
Small cladocerans, found abundantly on surfaces of macrophytes and sediments in the littoral zone, are important prey not only for small fish but also for various invertebrates such as larvae of ...odonates in freshwater habitats. However, no study has documented how habitat substrates affect their behavior and vulnerability to predators and predation. We conducted laboratory experiments to examine the movement of 3 littoral cladoceran species, Chydorus sphaericus, Alona sp., and Ilyocryptus spinifer, to determine if their vulnerabilities to predation by odonate larvae changed depending on the presence or absence of bottom sediment. We observed that when sediment was present, Ilyocryptus crawled in and ceased movement. However, in the containers without sediment, they continuously swam or crawled. Similarly, Chydorus also reduced frequency of movement in a container with sediment, but Alona movement did not change regardless of the presence or absence of sediment. In the predation experiments with 2 or 3 prey species, Ilyocryptus was the most vulnerable to predation by odonate larvae in the containers without sediment but least vulnerable in those with sediment. The vulnerability of Chydorus to the odonate larvae was as low as that of Ilyocryptus in the containers with sediment. Alona was less preyed upon by odonates in containers with sediment but highly vulnerable to predation when containers had sediment with Chydorus and Ilyocryptus. These results indicate that behavior and vulnerability to predation of littoral cladocerans are species-specific and change depending on the presence of sediment and the existence of other species.
Catalytic promiscuity of enzymes plays a pivotal role in driving the evolution of plant specialized metabolism. Chalcone synthase (CHS) catalyzes the production of 2′,4,4′,6′-tetrahydroxychalcone ...(THC), a common precursor of plant flavonoids, from p-coumaroyl-coenzyme A (-CoA) and three malonyl-CoA molecules. CHS has promiscuous product specificity, producing a significant amount of p-coumaroyltriacetic lactone (CTAL) in vitro. However, mechanistic aspects of this CHS promiscuity remain to be clarified. Here, we show that the product specificity of soybean CHS (GmCHS1) is altered by CoA, a reaction product, which selectively inhibits THC production (IC50, 67 μM) and enhances CTAL production. We determined the structure of a ternary GmCHS1/CoA/naringenin complex, in which CoA is bound to the CoA-binding tunnel via interactions with Lys55, Arg58, and Lys268. Replacement of these residues by alanine resulted in an enhanced THC/CTAL production ratio, suggesting the role of these residues in the CoA-mediated alteration of product specificity. In the ternary complex, a mobile loop (“the K-loop”), which contains Lys268, was in a “closed conformation” placing over the CoA-binding tunnel, whereas in the apo and binary complex structures, the K-loop was in an “open conformation” and remote from the tunnel. We propose that the production of THC involves a transition of the K-loop conformation between the open and closed states, whereas synthesis of CTAL is independent of it. In the presence of CoA, an enzyme conformer with the closed K-loop conformation becomes increasingly dominant, hampering the transition of K-loop conformations to result in decreased THC production and increased CTAL production.
•Catalytic promiscuity of enzymes drives evolution of plant specialized metabolism.•Chalcone synthase shows promiscuous product specificity, influenced by CoA.•Crystallographic studies were done to clarify the chalcone synthase promiscuity.•Conformational transition of “K-loop” during chalcone synthase catalysis is proposed.•CoA's impact on promiscuous product specificity is explored through K-loop dynamics.
Mushrooms are a favourite natural food in many countries. However, some wild species cause food poisoning, sometimes lethal, due to misidentification caused by confusing fruiting bodies similar to ...those of edible species. The morphological inspection of mycelia, spores and fruiting bodies have been traditionally used for the identification of mushrooms. More recently, DNA sequencing analysis has been successfully applied to mushrooms and to many other species. This study focuses on a simpler and more rapid methodology for the identification of wild mushrooms via protein profiling based on matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF MS). A preliminary study using 6 commercially available cultivated mushrooms suggested that a more reproducible spectrum was obtained from a portion of the cap than from the stem of a fruiting body by the extraction of proteins with a formic acid-acetonitrile mixture (1 + 1). We used 157 wild mushroom-fruiting bodies collected in the centre of Hokkaido from June to November 2014. Sequencing analysis of a portion of the ribosomal RNA gene provided 134 identifications of mushrooms by genus or species, however 23 samples containing 10 unknown species that had lower concordance rate of the nucleotide sequences in a BLAST search (less than 97%) and 13 samples that had unidentifiable poor or mixed sequencing signals remained unknown. MALDI-TOF MS analysis yielded a reproducible spectrum (frequency of matching score ≥ 2.0 was ≥6 spectra from 12 spectra measurements) for 114 of 157 samples. Profiling scores that matched each other within the database gave correct species identification (with scores of ≥2.0) for 110 samples (96%). An in-house prepared database was constructed from 106 independent species, except for overlapping identifications. We used 48 wild mushrooms that were collected in autumn 2015 to validate the in-house database. As a result, 21 mushrooms were identified at the species level with scores ≥2.0 and 5 mushrooms at the genus level with scores ≥1.7, although the signals of 2 mushrooms were insufficient for analysis. The remaining 20 samples were recognized as “unreliable identification” with scores <1.7. Subsequent DNA analysis confirmed that the correct species or genus identifications were achieved by MALDI-TOF MS for the 26 former samples, whereas the 18 mushrooms with poorly matched scores were species that were not included in the database. Thus, the proposed MALDI-TOF MS coupled with our database could be a powerful tool for the rapid and reliable identification of mushrooms; however, continuous updating of the database is necessary to enrich it with more abundant species.
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•An alternative method for species identification of wild mushroom was proposed.•Rapid and reliable identification was achieved by use of MALDI-TOF MS.•The lamella portion of the fruiting body provided the best spectrum signal.•An in-house database for wild mushroom was constructed.•More than 90% of the wild mushrooms collected in the next season were correctly inspected.
White bamboo is the swollen stem of Zizania latifolia parasitized by the smut fungus Ustilago esculenta. Five samples of galls were harvested at different stages of swelling, along with young ...seedlings of Z. latifolia and isolated colonies of U. esculenta. The inhibition capacity of boiling water or 50% ethanol extracts on NO release by RAW264 cells, which was stimulated with lipopolysaccharide, was evaluated as anti-inflammatory activity, while the NO induction capacity was evaluated as immunostimulatory activity. Total polyphenol (TPP) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging activity were also measured. The anti-inflammatory effect, as well as TPP and DPPH, was comprehensively detected in the ethanol extracts. The immunostimulating activity was observed in the boiled extracts at different levels, depending on the swelling stage, and was especially high in the top (apical) part. These data may indicate that functional components could be dynamically induced by interactions between Z. latifolia and U. esculenta.
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