Influence of pasteurization and spray drying on the fat digestion behavior of human milk fat analog emulsion: a simulated in vitro infant digestion study.

BACKGROUND: Human milk fat analog emulsion (HMFAE) is an emulsion that mimics the composition and structure of human milk (HM) fat globules. The application of HMFAE in infant formula requires a series of milk powder processing steps, such as pasteurization and spray drying. However, the effect of milk powder processing on fat digestion of HMFAE is still unclear. In this study, the influence of pasteurization and spray drying on the lipolysis behavior of HMFAE was studied and compared with HM using a simulated infant in vitro digestion model. RESULTS: Pasteurization and spray drying increased the flocculation and aggregation of lipid droplets in HMFAE during digestion. Spray drying destroyed the lipid droplet structure of HMFAE, and partial milk fat globule membrane-covered lipid droplets turned into protein-covered lipid droplets, which aggravated lipid-protein aggregation during gastric digestion and hindered fat digestion in the small intestine. The final lipolysis degree was in the order HM (64.55%) > HMFAE (63.41%) > pasteurized HMFAE (61.75%) > spray-dried HMFAE (60.57%). After complete gastrointestinal digestion, there were no significant differences in free fatty acid and sn-2 monoacylglycerol profile among the HMFAE, pasteurized HMFAE, and spray-dried HMFAE. CONCLUSION: Milk powder processing can reduce lipolysis by altering the lipid droplet structure of HMFAE and the degree of lipid droplet aggregation during digestion. © 2024 Society of Chemical Industry.

Wound size and location affect the prognosis of penetrating ocular injury.

BACKGROUND: Ocular trauma is a leading cause of vision loss. Penetrating ocular injury is a major type of open globe injury(OGI), while its epidemiology and clinical characteristics are still uncertain. The aim of this study is to reveal the prevalence and prognostic factors of penetrating ocular injury in the Shandong province. METHODS: A retrospective study of penetrating ocular injury was performed at the Second Hospital of Shandong University, from January 2010 to December 2019. Demographic information, injury causes, ocular trauma types, and initial and final visual acuity(VA) were analyzed. To obtain more precise characteristics of penetrating injury, the eye global was divided into three zones and analyzed. RESULTS: Among 210 OGI, there are 83 penetrating injuries which account for 39.5% of all. In addition, the final VA of 59 penetrating injuries recovered to 0.1 or better, which possesses the highest frequency among OGI. In order to research the relationship between the wound location and the final VA, we took 74 cases of penetrating injuries without retina or optic nerve damage for analysis. Results show that 62 were male and 12 were female. The average age was 36.01 ± 14.15. The most frequent occupation is the worker followed by the peasant. Statistics show that there is an obvious deviation in the Ocular trauma score (OTS) predicting the final VA and the actual final VA in the 45-65 score group (p < 0.05). Results suggest that the commonest penetrating injury zone is zone III (32 cases, 43.8%). Zone III, which is farthest from the center of the visual axis, has the largest improvement of the final VA (p = 0.0001). On the contrary, there is no statistical difference in the visual improvement in zone I and zone I + II that involves the injury of the central visual axis. CONCLUSION: This study describes the epidemiology and clinical characteristics of patients hospitalized for penetrating ocular injury without retina damage in Shandong province. It can be concluded that larger size and closer location to the visual axis of damage are accompanied by worse prognosis improvement. The study provides a better understanding of the disease and enlightenment for the prediction of visual prognosis.

Interfacial compositions of fat globules modulate structural characteristics and lipolysis of its model emulsions during in-vitro digestion.

BACKGROUND: This study investigated whether milk fat globule membrane as an emulsifier could make fat easier for infants to digest. An emulsion was formed using the membrane material, where anhydrous milk fat was used as the core material, milk fat globule membrane polar lipid (MPL) as the emulsifier, and soybean phospholipid (PL) and milk protein concentrate (MPC) incorporated as control emulsifiers. Structural characterization, glyceride composition, and fatty acid release from emulsions by in vitro digestion were investigated. RESULTS: The average particle size at the end of intestinal digestion was in the order MPL < PL < MPC, with diameters of 3.41 ± 0.51 µm, 3.53 ± 0.47 µm, and 10.46 ± 2.33 µm respectively. Meanwhile, laser scanning confocal microscopy results also illustrated that MPL could reduce the degree of aggregation during digestion. The lipolysis degree of MPL emulsion was higher than that of PL and MPC emulsions. MPL not only released higher levels of long-chain fatty acids, such as C18:1, C18:2, C18:3, which are of great significance for infant growth and development, but also released increased levels of C20:4 (arachidonic acid) and C22:6 (docosahexaenoic acid) than PL and MPC emulsions did. CONCLUSION: Fat droplets enveloped by milk fat globule MPLs were easier to digest and are therefore more suitable for infant formula. © 2023 Society of Chemical Industry.

Effects of lactoferrin on intestinal flora of metabolic disorder mice.

To study the mechanism of lactoferrin (LF) regulating metabolic disorders in nutritionally obese mice through intestinal microflora. Twenty-one male C57BL/6 mice were randomly divided into 3 groups: control group, model group and LF treatment group. The mice in control group were fed with maintenance diet and drank freely. The mice in model group were fed with high fat diet and drank freely. The mice in LF treatment group were fed with high fat diet and drinking water containing 2% LF freely. Body weight was recorded every week. Visceral fat ratio was measured at week 12. Blood glucose and serum lipid level were detected by automatic biochemical analyzer. The gut microbiota of mice was examined using 16 s rRNA sequencing method. LF treatment significantly reduced the levels of visceral adipose ratio, blood glucose, triglyceride, total cholesterol and low-density lipoprotein cholesterol (LDL-C) in high-fat diet mice (p < 0.05). It can be seen that drinking water with 2% LF had a significant impact on metabolic disorders. At the same time, the Firmicutes/Bacteroidetes ratio(F/B) of LF treated mice was decreased. The abundance of Deferribacteres, Oscillibacter, Butyricicoccus, Acinetobacter and Mucispirillum in LF treatment group were significantly decreased, and the abundance of Dubosiella was significantly increased (p < 0.05). In the LF-treated group, the expression levels of glucose metabolism genes in gut microbiota were increased, and the expression levels of pyruvate metabolism genes were decreased. It can be seen that metabolic disorders were related to intestinal flora. In conclusion, LF regulates metabolic disorders by regulating intestinal flora.

Design of a chimaeric antigen and its use in the detection of IgG antibodies against rubella virus.

BACKGROUND: Rubella virus (RV) is the causative agent of rubella or German measles. Although most infections cause only mild self-limited measles-like illness, the infection in pregnant women can cause severe foetal malformation or even miscarriage, especially in the first 3 months of pregnancy. Therefore, it is of great practical significance to establish a simple and sensitive RV detection method. METHODS: The partial epitopes of the E1 and E2 proteins from Rubella Virus were selected as the target sites, the sequence of the selected antigenic sites of the E1 and E2 were linked by a linker. The expression plasmid P6T was constructed by inserting the gene into PET-32A + with a histidine Tag. The P6 protein was induced and expressed in Escherichia coli L21 (DE3) and purified by nickel column affinity. The protein P6 antigen was identified by Western blotting analysis, and an anti-P6 antibody ELISA was established to test known serum samples to evaluate the capability of this method. RESULTS: After purification, the concentration and purity of the protein P6 were 0.283 mg/mL and more than 80%, respectively. Western blotting analysis showed that the protein P6 could react with rubella virus positive serum. By ELISA, 36 negative sera and 58 positive sera were detected. The coincidence rate, specificity and sensitivity of the ELISA were 86.2%, 88.89% and 84.48%, respectively. The P6 ELISA with a kappa coefficient of 0.715, P < 0.05, indicated excellent consistency. CONCLUSIONS: The protein P6 with excellent antigenicity obtained from prokaryotic expression followed by chromatography purification could prove useful for early diagnosis of RV infection.

Oral administration of Lactobacillus brevis 23017 combined with ellagic acid attenuates intestinal inflammatory injury caused by Eimeria infection by activating the Nrf2/HO-1 antioxidant pathway.

The aim of this study was to investigate whether oral administration of Lactobacillus brevis 23017 (LB) alone and in combination with ellagic acid inhibits ChTLR15/ChNLRP3/ChIL-1ß by activating the Nrf2/HO-1 pathway to attenuate intestinal inflammatory injury. Two animal experiments were performed. In Experiment 1, chickens were allocated into 7 groups: PBS, and low, medium and high dosages of live and heat-killed LB, named L/LB(+), M/LB(+) and H/LB(+), and L/LB(-), M/LB(-) and H/LB(-), respectively. In Experiment 2, chickens were divided into 5 groups: PBS, challenge control, and low, medium and high dosages of ellagic acid combined with LB(+), named L/EA + L/LB(+), M/EA + M/LB(+) and H/EA + H/LB(+), respectively. Chickens were gavaged with LB with or without ellagic acid once a day. Then, the mRNA and protein levels of the components of the Nrf2/HO-1 pathway found in the caecal tissues were quantified. On Day 7 post-infection with E. tenella, the levels of the components of the ChTLR15/NLRP3/IL-1ß pathway in the caeca were again quantified, and the anticoccidial effects were assessed. The results showed that the levels of the genes in the Nrf2/HO-1 pathway in the chickens in the LB(+) groups were higher than those in the LB(-) groups (p < 0.001); those in the H/LB(+) group were higher than those in the M/LB(+) and L/LB(+) groups (p < 0.001); and those in the H/EA + H/LB(+) group showed the highest expression levels compared with the other groups (p < 0.001). After challenge, the chickens in the H/LB(+) group displayed less inflammatory injury than those in the M/LB(+) and L/LB(+) groups (p < 0.05), and the chickens in the H/EA + H/LB(+) group showed stronger anti-inflammatory effects than the other groups (p < 0.05). Thus, these protective effects against infection were consistent with the above results. Overall, significant anti-inflammatory effects were observed in chickens orally gavaged with high dosages of live L. brevis 23017 and ellagic acid, which occurred by regulation of the ChTLR15/NLRP3/IL-1ß pathway.

Activation of ChTLR15/ChNF-κB-ChNLRP3/ChIL-1ß signaling transduction pathway mediated inflammatory responses to E. tenella infection.

Avian coccidiosis caused by Eimeria leads to severe economic losses in the global poultry industry. Although chicken Toll-like receptor 15 (ChTLR15) was reported to be involved in Eimeria infection, the detailed mechanism underlying its role in the inflammatory response remains to be discovered. The present study demonstrated that the mRNA expression levels of ChTLR15, ChMyD88, ChNF-κB, ChNLRP3, ChCaspase-1, ChIL-18 and ChIL-1ß and the protein levels of ChTLR15 and ChNLRP3 in cecal tissues of Eimeria-infected chickens were significantly elevated at 4, 12, and 24 h compared with those in noninfected control chickens (p < 0.01). Moreover, the mRNA levels of molecules in the ChTLR15/ChNF-κB and ChNLRP3/ChIL-1ß pathways and the protein levels of ChTLR15 and ChNLRP3 in chicken embryo fibroblast cells (DF-1) stimulated by E. tenella sporozoites were consistent with those in Eimeria-infected chickens. Furthermore, overexpression of ChTLR15 in DF1 cells augmented activation of the ChTLR15/ChNF-κB and ChNLRP3/ChIL-1ß pathways when stimulated with E. tenella sporozoites, while knockdown of ChTLR15 in DF1 cells showed inverse effects. Taken together, the present study provides evidence that E. tenella sporozoites specifically activate ChTLR15 and then trigger activation of the ChNLRP3/ChIL-1ß pathway, which partially mediates inflammatory responses to Eimeria infection.

Specific EtMIC3-binding peptides inhibit Eimeria tenella sporozoites entry into host cells.

Avian coccidiosis caused by Eimeria leads to huge economic losses on the global poultry industry. In this study, microneme adhesive repeat regions (MARR) bc1 of E. tenella microneme protein 3 (EtMIC3-bc1) was used as ligand, and peptides binding to EtMIC3 were screened from a phage display peptide library. The positive phage clones were checked by enzyme-linked immunosorbent assay (ELISA). Competitive ELISA was applied to further verify the binding capability between the positive phages and recombinant EtMIC3-bc1 protein or sporozoites protein. The inhibitory effects of target peptides on sporozoites invasion of MDBK cells were measured in vitro. Chickens were orally administrated with target positive phages and the protective effects against homologous challenge were evaluated. The model of three-dimensional (3D) structure for EtMIC3-bc1 was conducted, and molecular docking between target peptides and EtMIC3-bc1 model was analyzed. The results demonstrated that three selected positive phages specifically bind to EtMIC3-bc1 protein. The three peptides A, D and W effectively inhibited invasion of MDBK cells by sporozoites, showing inhibited ratio of 71.8%, 54.6% and 20.8%, respectively. Chickens in the group orally inoculated with phages A displayed more protective efficacies against homologous challenge than other groups. Molecular docking showed that amino acids in three peptides, especially in peptide A, insert into the hydrophobic groove of EtMIC3-bc1 protein, and bind to EtMIC3-bc1 through intermolecular hydrogen bonds. Taken together, the results suggest EtMIC3-binding peptides inhibit sporozoites entry into host cells. This study provides new idea for exploring novel strategies against coccidiosis.

Natural intertypic and intratypic recombinants of enterovirus 71 from mainland China during 2009-2018: a complete genome analysis.

Surveillance of recombinant enterovirus 71 (EV71) and subgenotype replacement is vital for preventing and controlling hand, foot, and mouth disease (HFMD) outbreaks. Despite this, data on recombinant variants and phylogeny of circulating EV71 strains in mainland China are limited. In this study, recombinant variants of EV71 were identified in mainland China from 2009 to 2018. Phylogenetic analysis indicated that except for individual strains (CQ2014-86/CQ/CHN/2014 and EV71/Xiamen/2009 (B5)), almost all of the EV71 strains in mainland China belonged to the subgenotype C4a. Analysing complete genome sequences of 196 EV71 isolates, 3 intertypic recombination strains (VR1432, 30-2/2015/BJ, and Guangdong-2009) and 5 intratypic recombination strains (EV71/P1034/2013, VR1432, Henan-ZMD/CHN/2012, Hubei-WH/CHN/2012, and EV71/P868/2013/China) were identified among naturally circulating EV71. The breakpoints of these recombinant strains were located within the P1, P2, and P3 encoding regions. Notably, a double recombinant (VR1432) resulting from recombination between EV71 subgenotype C4a and C4b strain SHZH98 and a CA8 strain Donovan was identified. This study reports these specific intertypic and intratypic recombination events for the first time highlighting the importance of genetic recombination in the emergence of new enterovirus variants.

Redescription and molecular characterization of sarcocysts of Sarcocystis cymruensis from Norway rats (Rattus norvegicus) and Sarcocystis ratti from black rats (R. rattus) in China.

In the present study, sarcocysts of Sarcocystis cymruensis were found in four of 42 (9.5%) Norway rats and those of S. ratti were observed in six of 60 (10%) black rats in China. With light microscopy, the sarcocysts of the two parasites were microscopic, and had smooth, thin cyst walls (≤ 1 µm). Ultrastructurally, the sarcocysts of S. cymruensis had small, osmiophilic, bleb-like protrusions, similar to type 1c; those of S. ratti had a cyst wall with regular, short, conical protrusions, similar to type 1 g. Three loci, i.e., 18S rDNA, the mitochondrial cox1 gene (Cox1), and the mitochondrial Cytb gene (Cytb), of the two parasites were sequenced and analyzed, and the Cytb sequences of the two parasites constituted the first records of this marker in GenBank. A comparison of the newly obtained sequences of the three loci between the two parasites revealed that the interspecific similarities of 18S rDNA, Cox1, and Cytb were 96.4-97.2%, 96.5%, and 93.7%, respectively. Therefore, the two species could be better discriminated with Cytb than with 18S rDNA and Cox1. Phylogenetic analysis based on 18S rDNA sequences and Cox1 sequences indicated that the two parasites had a close relationship with Sarcocystis in nonruminant animals, especially birds and canids.

A FRET-based ratiometric two-photon fluorescent probe for superoxide anion detection and imaging in living cells and tissues.

The superoxide anion (O2˙-) plays a crucial role in several physiological processes and many human diseases. Developing new methods for O2˙- detection in biological systems is very important. A FRET-based two-photon (TP) fluorescent probe with a ratiometric signal, TFR-O, was developed. A naphthalene derivative based TP fluorescent group was selected as the energy donor group, and a rhodol fluorescent group was chosen as the energy acceptor; the trifluoromethanesulfonate group was chosen as the recognition moiety. After reacting with O2˙-, the recognition moiety was removed and the fluorophore was released, leading to a fluorescence intensity decrease at the wavelength of 425 nm and a significant enhancement of the fluorescence intensity at 550 nm. The fluorescence intensity ratio between 550 and 425 nm (I550/I425) varied from 0.15 to 6.72, with the O2˙- concentration increasing from 0 to 50 µM. The detection limit of the TFR-O was 83 nM. Moreover, TFR-O was applied for detecting and imaging O2˙- in cells and liver tissues.

Correlations of severity of asthma in children with body mass index, adiponectin and leptin.

BACKGROUND: This study aims to investigate the correlations of asthma in children with body mass index (BMI), adiponectin, and leptin. METHODS: A total of 122 children with asthma in our hospital from January 2017 to February 2018 were randomly selected and divided into control group (normal) and observation group (BMI > 28 kg/m2 ) according to BMI. BMI, adiponectin, and leptin levels between the two groups were measured and compared, and correlations of disease grade with BMI, adiponectin, or leptin were analyzed. Moreover, risk factors for asthma in children were also identified. RESULTS: Body mass index, leptin level, forced vital capacity (FVC), FVC%, and forced expiratory volume in 1s (FEV1)/FVC in observation group were significantly higher than those in control group (P < 0.05), while the adiponectin level, forced expiratory capacity in 1s (FEC1), and FEV1% in observation group were significantly lower than those in control group (P < 0.05). The amount of severe patients in observation group was much larger than that in control group. The severity of disease was positively correlated with BMI and leptin and negatively correlated with adiponectin. BMI, adiponectin, and leptin were identified as risk factors for asthma in children. CONCLUSION: Adiponectin, leptin, and BMI are involved in the pathogenesis of asthma in children, suggesting they might be therapeutic targets for clinical treatment.

CNGC2 Is a Ca2+ Influx Channel That Prevents Accumulation of Apoplastic Ca2+ in the Leaf.

Ca2+ is absorbed by roots and transported upward through the xylem to the apoplastic space of the leaf, after which it is deposited into the leaf cell. In Arabidopsis (Arabidopsis thaliana), the tonoplast-localized Ca2+/H+ transporters CATION EXCHANGER1 (CAX1) and CAX3 sequester Ca2+ from the cytosol into the vacuole, but it is not known what transporter mediates the initial Ca2+ influx from the apoplast to the cytosol. Here, we report that Arabidopsis CYCLIC NUCLEOTIDE-GATED CHANNEL2 (CNGC2) encodes a protein with Ca2+ influx channel activity and is expressed in the leaf areas surrounding the free endings of minor veins, which is the primary site for Ca2+ unloading from the vasculature and influx into leaf cells. Under hydroponic growth conditions, with 0.1 mm Ca2+, both Arabidopsis cngc2 and cax1cax3 loss-of-function mutants grew normally. Increasing the Ca2+ concentration to 10 mm induced H2O2 accumulation, cell death, and leaf senescence and partially suppressed the hypersensitive response to avirulent pathogens in the mutants but not in the wild type. In vivo apoplastic Ca2+ overaccumulation was found in the leaves of cngc2 and cax1cax3 but not the wild type under the 10 mm Ca2+ condition, as monitored by Oregon Green BAPTA 488 5N, a low-affinity and membrane-impermeable Ca2+ probe. Our results indicate that CNGC2 likely has no direct roles in leaf development or the hypersensitive response but, instead, that CNGC2 could mediate Ca2+ influx into leaf cells. Finally, the in vivo extracellular Ca2+ imaging method developed in this study provides a new tool for investigating Ca2+ dynamics in plant cells.

Recombinant lactococcus lactis expressing Eimeria tenella AMA1 protein and its immunological effects against homologous challenge.

Avian coccidiosis leads to severe economic losses for the global poultry industry. Apical membrane antigen 1 (AMA1) of E. tenella (EtAMA1) plays a vital role during invasion of parasites into host cells. In the present study, recombinant live Lactococcus lactis expressing cytoplasmic, secreted and cell wall-anchored EtAMA1 protein were respectively constructed. The three live bacteria were respectively administered orally to SPF chickens (100 µl bacteria containing 5 × 109 CFU per chicken) for three times at 10-day intervals. After immunization, the lymphocyte proliferative function, the percentage of CD4+ and CD8α+ T cells in peripheral blood, and the IgG titers in serum of chickens in each group were respectively measured. The protective effects of live bacteria expressing EtAMA1 protein against E. tenella challenge were evaluated based on body weight gain (BWG), lesion score in cecum, oocyst descrease ratio. The results showed that chickens immunized with three live bacteria, especially the bacteria expressing cell wall-anchored EtAMA1 protein, displayed higher IgG titers and CD4+ T cells proportions, thus provided more immune protective effects against homologous challenge compared with the PBS control group and vector control group (lactococci harboring pTX8048). The oocyst decrease ratio of 33.33% from chickens immunized with lactococci expressing cell wall-anchored EctoAMA1 was observed, which was higher than that of 27.67% and 25.37% from the other two bacteria-immunized groups, respectively. The above results suggested that cell wall-anchored EtAMA1 protein delivered by Lactococcus lactis could stimulate an effective immune responses against Eimeria infection.

LncRNA Gm15290 sponges miR-27b to promote PPARγ-induced fat deposition and contribute to body weight gain in mice.

We found that Gm15290 was one of the most upregulated lncRNAs in the adipose of ob/ob mice through lncRNA microarray analysis. Then, manipulations of overexpression and silencing in mouse primary adipocytes showed that Gm15290 positively regulated adipogenesis, manifested by increasing lipid deposition and upregulating adipogenic genes including PPARγ, C/EBPα, and aP2. However, overexpression of mutant Gm15290 (at the binding site of miR-27c) did not have an promoting effect on adipogenesis. Additionally, Gm15290 was found to potentially interact with miR-27b that had been identified as a PPARγ targeting miRNA, and we verified their interaction by luciferase activity and RNA pull down assays. Furthermore, inhibition of Gm15290, by injection of the Gm15290 siRNA, decreased the body weight gain and mass of adipose tissues, including iWAT and eWAT, in mice fed with HFD. In conclusion, Gm15290 sponges miR-27b to increase fat deposition and body weight in HFD-fed mice.

Construction of Lactococcus lactis expressing secreted and anchored Eimeria tenella 3-1E protein and comparison of protective immunity against homologous challenge.

Two novel plasmids pTX8048-SP-Δ3-1E and pTX8048-SP-NAΔ3-1E-CWA were constructed. The plasmids were respectively electrotransformed into L. lactis NZ9000 to generate strain of L. lactis/pTX8048-SP-Δ3-1E in which 3-1E protein was expressed in secretion, and L. lactis/pTX8048-SP-NAΔ3-1E-CWA on which 3-1E protein was covalently anchored to the surface of bacteria cells. The expression of target proteins were examined by Western blot. The live lactococci expressing secreted 3-1E protein, anchored 3-1E protein, and cytoplasmic 3-1E protein was administered orally to chickens respectively, and the protective immunity and efficacy were compared by animal experiment. The results showed oral immunization to chickens with recombinant lactococci expressing anchored 3-1E protein elicited high 3-1E-specific serum IgG, increased high proportion of CD4+ and CD8α+ cells in spleen, alleviated average lesion score in cecum, decreased the oocyst output per chicken compared to lactococci expressing cytoplasmic or secreted 3-1E protein. Taken together, these findings indicated the surface anchored Eimeria protein displayed by L. lacits can induce protective immunity and partial protection against homologous infection.

Arabidopsis ROOT HAIR DEFECTIVE3 is involved in nitrogen starvation-induced anthocyanin accumulation.

Anthocyanin accumulation is a common phenomenon seen in plants under environmental stress. In this study, we identified a new allele of ROOT HAIR DEFECTIVE3 (RHD3) showing an anthocyanin overaccumulation phenotype under nitrogen starvation conditions. It is known that ethylene negatively regulates light- and sucrose-induced anthocyanin biosynthesis. We hypothesized that RHD3 achieves its negative effect on anthocyanin biosynthesis via an ethylene-regulating pathway. In support of this, similar to rhd3 mutants, the Arabidopsis ethylene signaling mutants etr1, ein2, and ein3/eil1 showed an anthocyanin overaccumulation phenotype under nitrogen starvation conditions. The ethylene precursor ACC strongly suppressed anthocyanin accumulation, dependent on ETR1, EIN2, EIN3/EIL1, and, partially, RHD3. In addition, inactivating RHD3 partially reversed the suppressive effect of ETO1 inactivation-evoked endogenous ethylene production on anthocyanin accumulation. The expression of nitrogen starvation-induced anthocyanin biosynthesis genes was negatively regulated by RHD3, but ethylene response genes were positively regulated by RHD3. Wild-type seedlings overexpressing RHD3 showed similar phenotypes to rhd3 mutants, indicating the existence of a fine-tuned relationship between gene expression and function. RHD3 was initially identified as a gene involved in root hair development. This study uncovered a new physiological function of RHD3 in nitrogen starvation-induced anthocyanin accumulation and ethylene homeostasis. [Correction added on 6 August 2015, after first online publication: "RND3" corrected to "RHD3".].

Glutamate receptor-like channel3.3 is involved in mediating glutathione-triggered cytosolic calcium transients, transcriptional changes, and innate immunity responses in Arabidopsis.

The tripeptide reduced glutathione (GSH; γ-glutamate [Glu]-cysteine [Cys]-glycine) is a major endogenous antioxidant in both animal and plant cells. It also functions as a neurotransmitter mediating communication among neurons in the central nervous system of animals through modulating specific ionotropic Glu receptors (GLRs) in the membrane. Little is known about such signaling roles in plant cells. Here, we report that transient rises in cytosolic calcium triggered by exogenous GSH in Arabidopsis (Arabidopsis thaliana) leaves were sensitive to GLR antagonists and abolished in loss-of-function atglr3.3 mutants. Like the GSH biosynthesis-defective mutant PHYTOALEXIN DEFICIENT2, atglr3.3 showed enhanced susceptibility to the bacterial pathogen Pseudomonas syringae pv tomato DC3000. Pathogen-induced defense marker gene expression was also decreased in atglr3.3 mutants. Twenty-seven percent of genes that were rapidly responsive to GSH treatment of seedlings were defense genes, most of which were dependent on functional AtGLR3.3, while GSH suppressed pathogen propagation through the AtGLR3.3-dependent pathway. Eight previously identified putative AtGLR3.3 ligands, GSH, oxidized glutathione, alanine, asparagine, Cys, Glu, glycine, and serine, all elicited the AtGLR3.3-dependent cytosolic calcium transients, but only GSH and Cys induced the defense response, with the Glu-induced AtGLR3.3-dependent transcription response being much less apparent than that triggered by GSH. Together, these observations suggest that AtGLR3.3 is required for several signaling effects mediated by extracellular GSH, even though these effects may not be causally related.

Pressure-driven variations of hydrogen bonding energy in ammonium azide (NH4N3): IR absorption and Raman scattering studies.

In this study, high pressure infrared (IR) absorption and Raman scattering studies for ammonium azide (NH4N3) were carried out at room temperature up to 20 GPa and 22 GPa, respectively. For comparison and further assignment, the vibrational spectra at ambient conditions were calculated using CASTEP code, particularly for the far- and mid-IR modes. The recorded vibrational data consistently indicated a pressure-induced phase transition at 2.9 GPa. All observed vibrational modes maintained their identities at the high pressure phase, indicating that NH4N3 was still presented in the form of ammonium cations and azide anions linked by the hydrogen bond (N-H⋯N). Above 2.9 GPa, the relative magnitude of the torsional mode weakened and the N-H symmetric stretch displayed a redshift, indicating strengthened hydrogen bonding energy. The opposite effects were observed above 12 GPa, where the relative magnitude of the torsional mode strengthened and the N-H symmetric stretch reverted to a blueshift, indicating weakened hydrogen bonding energy. It can be concluded that the hydrogen bonding energy exhibited a weakening (0-2.9 GPa), strengthening (2.9-12 GPa), and then again weakening (12-22 GPa) phenomena with the increasing of compression. The hydrogen bonding energy changing with the increase of pressure can be ascribed to a phase transition at 2.9 GPa and a rotational or bending behavior of azide ions at 12 GPa.

AtPEPTIDE RECEPTOR2 mediates the AtPEPTIDE1-induced cytosolic Ca(2+) rise, which is required for the suppression of Glutamine Dumper gene expression in Arabidopsis roots.

AtPEPTIDE RECEPTOR2 (AtPEPR2) is a member of leucine-rich repeat receptor-like kinase family and binds to a group of AtPROPEP gene-encoded endogenous peptides, AtPeps. Previously, we found that AtPEPR2 plays a moderate role in the AtPep1-mediated innate immunity responses in Arabidopsis leaf. In this study, we found that AtPEPR2 promoter has strong activity in the vascular tissues of the roots and the atpepr2 mutants showed a moderate but significantly shorter root phenotype. AtPEPR2 partially mediated AtPep1-induced root elongation inhibition. AtPep1-triggered cytosolic Ca(2+) transient rise in roots showed partial dependence on AtPEPR2 and fully on extracellular Ca(2+) ([Ca(2+) ]ext ). Transcriptional profiling analysis found that expression of 75% of AtPep1-modulated genes in roots was fully dependent on AtPEPR2, of which two dramatically induced genes showed partial dependence on the [Ca(2+) ]ext . Arabidopsis genome contains seven Glutamine Dumpers genes (AtGDUs), encoding amino acid exporters. Three of them (AtGDU2, 3, 5) were among the top 10 genes that were downregulated by AtPep1 through AtPEPR2 fully dependent pathway. Treatment with AtPep1 strongly suppressed promoter activity of AtGDU3 in roots, which was relieved by chelating [Ca(2+) ]ext . Arabidopsis overexpressing AtGDU3 showed a shorter root phenotype and decreased sensitivity to the AtPep1-mediated inhibition of root elongation. Taken together, this study demonstrated a significant role of AtPEPR2 in the AtPep1-mediated signaling in the roots.