Wet-type grinder-treated okara modulates gut microbiota composition and attenuates obesity in high-fat-fed mice.

Wet-type grinder (WG) is a nanofiber technology used to atomize dietary fiber-rich materials. WG-treated okara (WGO) exhibits high dispersion and viscosity similar to those of viscous soluble dietary fibers. Here, we studied the effect of WGO supplementation on obesity and gut microbiota composition in high-fat diet (HFD)-fed mice. WGO intake suppressed body weight gain and fat accumulation, improved glucose tolerance, lowered cholesterol levels, and prevented HFD-induced decrease in muscle mass. WGO supplementation also led to cecum enlargement, lower pH, and higher butyrate production. The bacterial 16S ribosomal RNA genes (16S rDNA) were sequenced to determine the gut microbiota composition of the fecal samples. Sequencing of bacterial 16S rDNA revealed that WGO treatment increased the abundance of butyrate producer Ruminococcus and reduced the abundances of Rikenellaceae, Streptococcaceae, and Prevotellaceae, which are related to metabolic diseases. Metabolomics analysis of the plasma of WGO- and cellulose-treated mice were conducted using ultra-high-performance liquid chromatography-mass spectrometry. Metabolic pathway analysis revealed that the primary bile acid biosynthesis pathway was significantly positively regulated by WGO intake instead of cellulose. These results demonstrate that WG is useful for improving functional properties of okara to prevent metabolic syndromes, including obesity, diabetes, and dyslipidemia.

The disordered C-terminal tail of fungal LPMOs from phytopathogens mediates protein dimerization and impacts plant penetration.

Lytic polysaccharide monooxygenases (LPMOs) are monocopper enzymes that oxidatively degrade various polysaccharides, such as cellulose. Despite extensive research on this class of enzymes, the role played by their C-terminal regions predicted to be intrinsically disordered (dCTR) has been overlooked. Here, we investigated the function of the dCTR of an LPMO, called CoAA9A, up-regulated during plant infection by Colletotrichum orbiculare, the causative agent of anthracnose. After recombinant production of the full-length protein, we found that the dCTR mediates CoAA9A dimerization in vitro, via a disulfide bridge, a hitherto-never-reported property that positively affects both binding and activity on cellulose. Using SAXS experiments, we show that the homodimer is in an extended conformation. In vivo, we demonstrate that gene deletion impairs formation of the infection-specialized cell called appressorium and delays penetration of the plant. Using immunochemistry, we show that the protein is a dimer not only in vitro but also in vivo when secreted by the appressorium. As these peculiar LPMOs are also found in other plant pathogens, our findings open up broad avenues for crop protection.

Palaeoproteomic investigation of an ancient human skeleton with abnormal deposition of dental calculus.

Detailed investigation of extremely severe pathological conditions in ancient human skeletons is important as it could shed light on the breadth of potential interactions between humans and disease etiologies in the past. Here, we applied palaeoproteomics to investigate an ancient human skeletal individual with severe oral pathology, focusing our research on bacterial pathogenic factors and host defense response. This female skeleton, from the Okhotsk period (i.e., fifth to thirteenth century) of Northern Japan, poses relevant amounts of abnormal dental calculus deposition and exhibits oral dysfunction due to severe periodontal disease. A shotgun mass-spectrometry analysis identified 81 human proteins and 15 bacterial proteins from the calculus of the subject. We identified two pathogenic or bioinvasive proteins originating from two of the three "red complex" bacteria, the core species associated with severe periodontal disease in modern humans, as well as two additional bioinvasive proteins of periodontal-associated bacteria. Moreover, we discovered defense response system-associated human proteins, although their proportion was mostly similar to those reported in ancient and modern human individuals with lower calculus deposition. These results suggest that the bacterial etiology was similar and the host defense response was not necessarily more intense in ancient individuals with significant amounts of abnormal dental calculus deposition.

Millmeter-wave irradiation regulates mRNA-expression and the ubiquitin-proteasome system in wheat exposed to flooding stress.

The irradiation with millimeter-wave (MMW) of wheat seeds promotes root growth under flooding stress; however, its role is not completely clarified. Nuclear proteomics was performed, to reveal the role of MMW irradiation in enhancing flooding tolerance. The purity of nuclear fractions purified from roots was verified. Histone, which is a protein marker for nuclear-purification efficiency, was enriched; and cytosolic ascorbate peroxidase was reduced in the nuclear fraction. The principal-component analysis of proteome displayed that the irradiation of seeds affected nuclear proteins in roots grown under flooding stress. Proteins detected using proteomic analysis were verified using immunoblot analysis. Histone H3 accumulated under flooding stress; however, it decreased to the control level by irradiation. Whereas the ubiquitin accumulated in roots grown under stress when seeds were irradiated. These results suggest that MMW irradiation improves wheat-root growth under flooding stress through the regulation of mRNA-expression level and the ubiquitin-proteasome system. SIGNIFICANCE: To reveal the role of millimeter-wave irradiation in enhancing flooding tolerance in wheat, nuclear proteomics was performed. The principal-component analysis of proteome displayed that irradiation of seeds affected nuclear proteins in roots grown under flooding stress. Proteins detected using proteomic analysis were verified using immunoblot analysis. Histone H3 accumulated under flooding stress; however, it decreased to the control level with irradiation. Whereas the ubiquitin accumulated in roots grown under stress when seeds were irradiated. These results suggest that millimeter-wave irradiation improves wheat-root growth under flooding stress through the regulation of mRNA-expression level and the ubiquitin-proteasome system.

Proteomic and molecular analyses to understand the promotive effect of safranal on soybean growth under salt stress.

Safranal is a free radical scavenger and useful as an antioxidant molecule; however, its promotive role in soybean is not explored. Salt stress decreased soybean growth and safranal improved it even if under salt stress. To study the positive mechanism of safranal on soybean growth, a proteomic approach was used. According to functional categorization, oppositely changed proteins were further confirmed using biochemical techniques. Actin and calcium-dependent protein kinase decreased in soybean root and hypocotyl, respectively, under salt stress and increased with safranal application. Xyloglucan endotransglucosylase/ hydrolase increased in soybean root under salt stress but decreased with safranal application. Peroxidase increased under salt stress and further enhanced by safranal application in soybean root. Actin, RuvB-like helicase, and protein kinase domain-containing protein were upregulated under salt stress and further enhanced by safranal application under salt stress. Dynamin GTPase was downregulated under salt stress but recovered with safranal application under salt stress. Glutathione peroxidase and PfkB domain-containing protein were upregulated by safranal application under salt stress in soybean root. These results suggest that safranal improves soybean growth through the regulation of cell wall and nuclear proteins along with reactive­oxygen species scavenging system. Furthermore, it might promote salt-stress tolerance through the regulation of membrane proteins involved in endocytosis and post-Golgi trafficking. SIGNIFICANCE: To study the positive mechanism of safranal on soybean growth, a proteomic approach was used. According to functional categorization, oppositely changed proteins were further confirmed using biochemical techniques. Actin and calcium-dependent protein kinase decreased in soybean root and hypocotyl, respectively, under salt stress and increased with safranal application. Xyloglucan endotransglucosylase/ hydrolase increased in soybean root under salt stress but decreased with safranal application. Peroxidase increased under salt stress and further enhanced by safranal application in soybean root. Actin, RuvB-like helicase, and protein kinase domain-containing protein were upregulated under salt stress and further enhanced by safranal application under salt stress. Dynamin GTPase was downregulated under salt stress but recovered with safranal application under salt stress. Glutathione peroxidase and PfkB domain-containing protein were upregulated by safranal application under salt stress in soybean root. These results suggest that safranal improves soybean growth through the regulation of cell wall and nuclear proteins along with reactive­oxygen species scavenging system. Furthermore, it might promote salt-stress tolerance through the regulation of membrane proteins involved in endocytosis and post-Golgi trafficking.

Identification of uromodulin deposition in the stroma of perinephric fibromyxoid nephrogenic adenoma by mass spectrometry.

Nephrogenic adenoma (NA) is an epithelial lesion that usually occurs in the mucosa of the urinary tract. Rare cases of deep infiltrative or perinephric lesions have also been reported. Recently, NA with characteristic fibromyxoid stroma (fibromyxoid NA) has been proposed as a distinct variant. Although shedding of distal renal tubular cells due to urinary tract rupture has been postulated as the cause of NA in general, the mechanism underlying extraurinary presentation of NA and fibromyxoid stromal change in fibromyxoid NA remains unknown. In this study, we performed mass spectrometry (MS) analysis in a case of perinephric fibromyxoid NA of an 82-year-old man who underwent right nephroureterectomy for distal ureteral cancer. The patient had no prior history of urinary tract injury or radiation. Periodic acid-Schiff staining-positive eosinophilic structureless deposits in the stroma of fibromyxoid NA were microdissected and subjected to liquid chromatography/MS. The analysis revealed the presence of a substantial amount of uromodulin (Tamm-Horsfall protein). The presence of urinary content in the stroma of perinephric fibromyxoid NA suggests that urinary tract rupture and engraftment of renal tubular epithelial cells directly cause the lesion.

Super-enhancer trapping by the nuclear pore via intrinsically disordered regions of proteins in squamous cell carcinoma cells.

Master transcription factors such as TP63 establish super-enhancers (SEs) to drive core transcriptional networks in cancer cells, yet the spatiotemporal regulation of SEs within the nucleus remains unknown. The nuclear pore complex (NPC) may tether SEs to the nuclear pore where RNA export rates are maximal. Here, we report that NUP153, a component of the NPC, anchors SEs to the NPC and enhances TP63 expression by maximizing mRNA export. This anchoring is mediated through protein-protein interaction between the intrinsically disordered regions (IDRs) of NUP153 and the coactivator BRD4. Silencing of NUP153 excludes SEs from the nuclear periphery, decreases TP63 expression, impairs cellular growth, and induces epidermal differentiation of squamous cell carcinoma. Overall, this work reveals the critical roles of NUP153 IDRs in the regulation of SE localization, thus providing insights into a new layer of gene regulation at the epigenomic and spatial level.

Multi-omics Analyses of Non-GM Tomato Scion Engrafted on GM Rootstocks.

Grafting has been widely applied in agricultural production in order to utilize agriculturally valuable traits. The use of genetically modified (GM) plants for grafting with non-GM crops will soon be implemented to generate chimeric plants (transgrafting)*, and the non-GM edible portions thus obtained could fall outside of the current legal regulations. A number of metabolites and macromolecules are reciprocally exchanged between scion and rootstock, affecting the crop properties as food. Accordingly, the potential risks associated with grafting, particularly those related to transgrafting with GM plants, should be carefully evaluated based on scientific evidence. In this study, we prepared a hetero-transgraft line composed of non-GM tomato scion and GM-tobacco rootstock expressing firefly luciferase. We also prepared a homograft line (both rootstock and scion are from non-GM tomato) and a heterograft line (non-GM tobacco rootstock and non-GM tomato scion). The non-GM tomato fruits were harvested from these grafted lines and subjected to comprehensive characterization by multi-omics analysis. Proteomic analysis detected tobacco-derived proteins from both heterograft and hetero-transgraft lines, suggesting protein transfer from the tobacco rootstock to the tomato fruits. No allergenicity information is available for these two tobacco-derived proteins. The transcript levels of the genes encoding two allergenic tomato intrinsic proteins (Sola l 4.0101 and Sola l 4.0201) decreased in the heterograft and hetero-transgraft lines. Several differences were observed in the metabolic profiles, including α-tomatine and nicotine. The accumulation of tobacco-derived nicotine in the tomato fruits of both heterograft and hetero-transgraft lines indicated that the transfer of unfavorable metabolites from rootstock to scion should be assessed as a food safety concern. Further investigations are needed to clarify whether variable environmental conditions and growth periods may influence the qualities of the non-GM edible parts produced by such transgrafted plants.

Renal Pharmacokinetic Adaptation to Cholestasis Causes Increased Nephrotoxic Drug Accumulation by Mrp6 Downregulation in Mice.

In hepatic dysfunction, renal pharmacokinetic adaptation can be observed, although information on the changes in drug exposure and the interorgan regulation of membrane transporters in kidney in liver diseases is limited. This study aimed to clarify the effects of renal exposure to nephrotoxic drugs during cholestasis induced by bile duct ligation (BDL). Among the 11 nephrotoxic drugs examined, the tissue accumulation of imatinib and cisplatin in kidney slices obtained from mice 2 weeks after BDL operation was higher than that in sham-operated mice. The uptake of imatinib in the kidney slices of BDL mice was slightly higher, whereas its efflux from the slices was largely decreased compared to that in sham-operated mice. Proteomic analysis revealed a reduction in renal expression of the efflux transporter multidrug resistance-associated protein 6 (Mrp6/Abcc6) in BDL mice, and both imatinib and cisplatin were identified as Mrp6 substrates. Survival probability after cisplatin administration was reduced in BDL mice. In conclusion, the present study demonstrated that BDL-induced cholestasis leads to the downregulation of the renal basolateral efflux transporter Mrp6, resulting in drug accumulation in renal cells and promoting drug-induced renal injury.

Cell assembly analysis of neural circuits for innate behavior in Drosophila melanogaster using an immediate early gene stripe/egr-1.

Innate behavior, such as courtship behavior, is controlled by a genetically defined set of neurons. To date, it remains challenging to visualize and artificially control the neural population that is active during innate behavior in a whole-brain scale. Immediate early genes (IEGs), whose expression is induced by neural activity, can serve as powerful tools to map neural activity in the animal brain. We screened for IEGs in vinegar fly Drosophila melanogaster and identified stripe/egr-1 as a potent neural activity marker. Focusing on male courtship as a model of innate behavior, we demonstrate that stripe-GAL4-mediated reporter expression can label fruitless (fru)-expressing neurons involved in courtship in an activity (experience)-dependent manner. Optogenetic reactivation of the labeled neurons elicited sexual behavior in males, whereas silencing of the labeled neurons suppressed courtship and copulation. Further, by combining stripe-GAL4-mediated reporter expression and detection of endogenous Stripe expression, we established methods that can label neurons activated under different contexts in separate time windows in the same animal. The cell assembly analysis of fru neural population in males revealed that distinct groups of neurons are activated during interactions with a female or another male. These methods will contribute to building a deeper understanding of neural circuit mechanisms underlying innate insect behavior.

Apple-derived extracellular vesicles modulate the expression of human intestinal bile acid transporter ASBT/SLC10A2 via downregulation of transcription factor RARα.

PURPOSE: Plant-derived extracellular vesicles (EVs) have been reported to exert biological activity on intestinal tissues by delivering their contents into intestinal cells. We previously reported that ASBT/SLC10A2 mRNA was downregulated by apple-derived extracellular vesicles (APEVs). ASBT downregulation is effective in the treatment of cholestasis and chronic constipation, similar to the beneficial effects of apples. Therefore, this study aimed to establish the mechanism of ASBT downregulation by APEVs, focusing on microRNAs present in APEVs. RESULTS: APEVs downregulated the expression of ASBT, but no significant effect on SLC10A2-3'UTR was observed. Proteomics revealed that APEVs decreased the expression of RARα/NR1B1. The binding of RARα to SLC10A2 promoter was also decreased by APEVs. The stability of NR1B1 mRNA was attenuated by APEVs and its 3'UTR was found to be a target for APEVs. Apple microRNAs that were predicted to interact with NR1B1-3'UTR were present in APEVs, and their mimics suppressed NR1B1 mRNA expression. CONCLUSIONS: Suppression of ASBT by APEVs was indirectly mediated by the downregulation of RARα, and its stability was lowered by microRNAs present in APEVs. This study suggested that macromolecules in food directly affect intestinal function by means of EVs that stabilize them and facilitate their cellular uptake.

Postnatal expression of CD38 in astrocytes regulates synapse formation and adult social memory.

Social behavior is essential for health, survival, and reproduction of animals; however, the role of astrocytes in social behavior remains largely unknown. The transmembrane protein CD38, which acts both as a receptor and ADP-ribosyl cyclase to produce cyclic ADP-ribose (cADPR) regulates social behaviors by promoting oxytocin release from hypothalamic neurons. CD38 is also abundantly expressed in astrocytes in the postnatal brain and is important for astroglial development. Here, we demonstrate that the astroglial-expressed CD38 plays an important role in social behavior during development. Selective deletion of CD38 in postnatal astrocytes, but not in adult astrocytes, impairs social memory without any other behavioral abnormalities. Morphological analysis shows that depletion of astroglial CD38 in the postnatal brain interferes with synapse formation in the medial prefrontal cortex (mPFC) and hippocampus. Moreover, astroglial CD38 expression promotes synaptogenesis of excitatory neurons by increasing the level of extracellular SPARCL1 (also known as Hevin), a synaptogenic protein. The release of SPARCL1 from astrocytes is regulated by CD38/cADPR/calcium signaling. These data demonstrate a novel developmental role of astrocytes in neural circuit formation and regulation of social behavior in adults.

Membrane Proteomics to Understand Enhancement Effects of Millimeter-Wave Irradiation on Wheat Root under Flooding Stress.

Millimeter-wave irradiation of wheat seeds enhances the growth of roots under flooding stress, but its mechanism is not clearly understood. To understand the role of millimeter-wave irradiation on root-growth enhancement, membrane proteomics was performed. Membrane fractions purified from wheat roots were evaluated for purity. H+-ATPase and calnexin, which are protein markers for membrane-purification efficiency, were enriched in a membrane fraction. A principal-component analysis of the proteomic results indicated that the millimeter-wave irradiation of seeds affects membrane proteins in grown roots. Proteins identified using proteomic analysis were confirmed using immunoblot or polymerase chain reaction analyses. The abundance of cellulose synthetase, which is a plasma-membrane protein, decreased under flooding stress; however, it increased with millimeter-wave irradiation. On the other hand, the abundance of calnexin and V-ATPase, which are proteins in the endoplasmic reticulum and vacuolar, increased under flooding stress; however, it decreased with millimeter-wave irradiation. Furthermore, NADH dehydrogenase, which is found in mitochondria membranes, was upregulated due to flooding stress but downregulated following millimeter-wave irradiation even under flooding stress. The ATP content showed a similar trend toward change in NADH dehydrogenase expression. These results suggest that millimeter-wave irradiation improves the root growth of wheat via the transitions of proteins in the plasma membrane, endoplasmic reticulum, vacuolar, and mitochondria.

Kynurenine promotes Calcitonin secretion and reduces cortisol in the Japanese flounder Paralichthys olivaceus.

Deep ocean water (DOW) exerts positive effects on the growth of marine organisms, suggesting the presence of unknown component(s) that facilitate their aquaculture. We observed that DOW suppressed plasma cortisol (i.e., a stress marker) concentration in Japanese flounder (Paralichthys olivaceus) reared under high-density condition. RNA-sequencing analysis of flounder brains showed that when compared to surface seawater (SSW)-reared fish, DOW-reared fish had lower expression of hypothalamic (i.e., corticotropin-releasing hormone) and pituitary (i.e., proopiomelanocortin, including adrenocorticotropic hormone) hormone-encoding genes. Moreover, DOW-mediated regulation of gene expression was linked to decreased blood cortisol concentration in DOW-reared fish. Our results indicate that DOW activated osteoblasts in fish scales and facilitated the production of Calcitonin, a hypocalcemic hormone that acts as an analgesic. We then provide evidence that the Calcitonin produced is involved in the regulatory network of genes controlling cortisol secretion. In addition, the indole component kynurenine was identified as the component responsible for osteoblast activation in DOW. Furthermore, kynurenine increased plasma Calcitonin concentrations in flounders reared under high-density condition, while it decreased plasma cortisol concentration. Taken together, we propose that kynurenine in DOW exerts a cortisol-reducing effect in flounders by facilitating Calcitonin production by osteoblasts in the scales.

An agroecological structure model of compost-soil-plant interactions for sustainable organic farming.

Compost is used worldwide as a soil conditioner for crops, but its functions have still been explored. Here, the omics profiles of carrots were investigated, as a root vegetable plant model, in a field amended with compost fermented with thermophilic Bacillaceae for growth and quality indices. Exposure to compost significantly increased the productivity, antioxidant activity, color, and taste of the carrot root and altered the soil bacterial composition with the levels of characteristic metabolites of the leaf, root, and soil. Based on the data, structural equation modeling (SEM) estimated that amino acids, antioxidant activity, flavonoids and/or carotenoids in plants were optimally linked by exposure to compost. The SEM of the soil estimated that the genus Paenibacillus and nitrogen compounds were optimally involved during exposure. These estimates did not show a contradiction between the whole genomic analysis of compost-derived Paenibacillus isolates and the bioactivity data, inferring the presence of a complex cascade of plant growth-promoting effects and modulation of the nitrogen cycle by the compost itself. These observations have provided information on the qualitative indicators of compost in complex soil-plant interactions and offer a new perspective for chemically independent sustainable agriculture through the efficient use of natural nitrogen.

Potential biomarker proteins for aspiration pneumonia detected by shotgun proteomics using buccal mucosa samples: a cross-sectional case-control study.

BACKGROUND: Aspiration pneumonia (AP), which is a major cause of death in the elderly, does present with typical symptoms in the early stages of onset, thus it is difficult to detect and treat at an early stage. In this study, we identified biomarkers that are useful for the detection of AP and focused on salivary proteins, which may be collected non-invasively. Because expectorating saliva is often difficult for elderly people, we collected salivary proteins from the buccal mucosa. METHODS: We collected samples from the buccal mucosa of six patients with AP and six control patients (no AP) in an acute-care hospital. Following protein precipitation using trichloroacetic acid and washing with acetone, the samples were analyzed by liquid chromatography and tandem mass spectrometry (LC-MS/MS). We also determined the levels of cytokines and chemokines in non-precipitated samples from buccal mucosa. RESULTS: Comparative quantitative analysis of LC-MS/MS spectra revealed 55 highly (P values < 0.10) abundant proteins with high FDR confidence (q values < 0.01) and high coverage (> 50%) in the AP group compared with the control group. Among the 55 proteins, the protein abundances of four proteins (protein S100-A7A, eukaryotic translation initiation factor 1, Serpin B4, and peptidoglycan recognition protein 1) in the AP group showed a negative correlation with the time post-onset; these proteins are promising AP biomarker candidates. In addition, the abundance of C-reactive protein (CRP) in oral samples was highly correlated with serum CRP levels, suggesting that oral CRP levels may be used as a surrogate to predict serum CRP in AP patients. A multiplex cytokine/chemokine assay revealed that MCP-1 tended to be low, indicating unresponsiveness of MCP-1 and its downstream immune pathways in AP. CONCLUSION: Our findings suggest that oral salivary proteins, which are obtained non-invasively, can be utilized for the detection of AP.

Omics Profiles of Non-GM Tubers from Transgrafted Potato with a GM Scion.

"Transgrafting" is a grafting procedure whereby a transgenic plant body is grafted to a non-transgenic plant body. It is a novel plant breeding technology that allows non-transgenic plants to obtain benefits usually conferred to transgenic plants. Many plants regulate flowering by perceiving the day-length cycle via expression of FLOWERING LOCUS T (FT) in the leaves. The resulting FT protein is translocated to the shoot apical meristem via the phloem. In potato plants, FT is involved in the promotion of tuber formation. Here we investigated the effects of a genetically modified (GM) scion on the edible parts of the non-GM rootstock by using potato plants transformed with StSP6A, a novel potato homolog of the FT gene. Scions prepared from GM or control (wild-type) potato plants were grafted to non-GM potato rootstocks; these were designated as TN and NN plants, respectively. After tuber harvest, we observed no significant differences in potato yield between TN and NN plants. Transcriptomic analysis revealed that only one gene-with unknown function-was differentially expressed between TN and NN plants. Subsequent proteomic analysis indicated that several members of protease inhibitor families, known as anti-nutritional factors in potato, were slightly more abundant in TN plants. Metabolomic analysis revealed a slight increase in metabolite abundance in NN plants, but we observed no difference in the accumulation of steroid glycoalkaloids, toxic metabolites found in potato. Finally, we found that TN and NN plants did not differ in nutrient composition. Taken together, these results indicate that FT expression in scions had a limited effect on the metabolism of non-transgenic potato tubers.

Tandem metalloenzymes gate plant cell entry by pathogenic fungi.

Global food security is endangered by fungal phytopathogens causing devastating crop production losses. Many of these pathogens use specialized appressoria cells to puncture plant cuticles. Here, we unveil a pair of alcohol oxidase-peroxidase enzymes to be essential for pathogenicity. Using Colletotrichum orbiculare, we show that the enzyme pair is cosecreted by the fungus early during plant penetration and that single and double mutants have impaired penetration ability. Molecular modeling, biochemical, and biophysical approaches revealed a fine-tuned interplay between these metalloenzymes, which oxidize plant cuticular long-chain alcohols into aldehydes. We show that the enzyme pair is involved in transcriptional regulation of genes necessary for host penetration. The identification of these infection-specific metalloenzymes opens new avenues on the role of wax-derived compounds and the design of oxidase-specific inhibitors for crop protection.

Reduced graphene oxide increases cells with enlarged outer membrane of Citrifermentans bremense and exopolysaccharides secretion.

Conductive carbons can boost anaerobic microbial metabolism by assisting extracellular electron transfer (EET), and their chemistry affects microbial metabolism. Graphene oxide (GO), a chemically oxidized sheet of graphite, has been used in various bioelectrochemical systems, although its mechanism is rarely understood. This study revealed specific metabolic responses to reduced GO (rGO) in an electrogenic strain R4 of Citrifermentans bremense, recently renamed from "Geobacter bremensis," in comparison to that with graphite felt (GF). Specifically, the change in growth from planktonic cells to biofilm with an enlarged outer membrane. The mRNA profile supported the fact that rGO upregulated the 14 genes related to the exopolysaccharides (EPS) secretion and biofilm formation, which is more than that in GF (4 genes). While GF upregulated the 35 genes involved in cell motility, which is more than that in rGO (8 genes). The heme protein profile suggested that both carbons induced similar EET pathways involving OmcA/MtrC and OmcS; however, GO specifically induced PilQ. These findings show that the chemistry of conductive carbon differentiates metabolism, especially affecting cellular morphology or living form, rather than electron transfer metabolism.

Proteomic Profiling of Plant and Pathogen Interaction on the Leaf Epidermis.

The plant epidermis is the first line of plant defense against pathogen invasion, and likely contains important regulatory proteins related to the plant-pathogen interaction. This study aims to identify the candidates of these regulatory proteins expressed in the plant epidermis. We performed comparative proteomic studies to identify rapidly and locally expressed proteins in the leaf epidermis inoculated with fungal phytopathogen. The conidia solutions were dropped onto the Arabidopsis leaf surface, and then, we collected the epidermal tissues from inoculated and mock-treated leaves at 4 and 24 hpi. The label-free quantification methods showed that expressions of Arabidopsis proteins, which are related to defense signals, such as BAK1, MKK5, receptor-like protein kinases, transcription factors, and stomatal functions, were rapidly induced in the epidermal tissues of inoculated leaves. In contrast, most of them were not differentially regulated by fugal inoculation in the whole leaves. These findings clearly indicate that epidermal proteomics can monitor locally expressed proteins in inoculated areas of plant tissues. We also identified the 61 fungal proteins, including effector-like proteins specifically expressed on the Arabidopsis epidermis. Our new findings suggested that epidermal proteomics is useful for understanding the local expressions of plant and fungal proteins related to their interactions.