Timely Questions Emerging in Chronobiology: The Circadian Clock Keeps on Ticking.

Chronobiology investigations have revealed much about cellular and physiological clockworks but we are far from having a complete mechanistic understanding of the physiological and ecological implications. Here we present some unresolved questions in circadian biology research as posed by the editorial staff and guest contributors to the Journal of Circadian Rhythms. This collection of ideas is not meant to be comprehensive but does reveal the breadth of our observations on emerging trends in chronobiology and circadian biology. It is amazing what could be achieved with various expected innovations in technologies, techniques, and mathematical tools that are being developed. We fully expect strengthening mechanistic work will be linked to health care and environmental understandings of circadian function. Now that most clock genes are known, linking these to physiological, metabolic, and developmental traits requires investigations from the single molecule to the terrestrial ecological scales. Real answers are expected for these questions over the next decade. Where are the circadian clocks at a cellular level? How are clocks coupled cellularly to generate organism level outcomes? How do communities of circadian organisms rhythmically interact with each other? In what way does the natural genetic variation in populations sculpt community behaviors? How will methods development for circadian research be used in disparate academic and commercial endeavors? These and other questions make it a very exciting time to be working as a chronobiologist.

Advanced treatment and Resource recovery of brewery wastewater by Co-cultivation of filamentous microalga Tribonema aequale and autochthonous Bacteria.

To use unicellular microalgae to remove waste nutrients from brewery wastewater while converting them into algal biomass has been explored but high-cost treatment and low-value biomass associated with current technologies have prevented this concept from further attempts. In this study, a filamentous microalga Tribonema aequale was introduced and the alga can grow vigorously in brewery wastewater and algal biomass concentration could be as high as 6.45 g L-1 which can be harvested by a cost-effective filtration method. The alga together with autochthonous bacteria removed majority of waste nutrients from brewery wastewater. Specifically, 85.39% total organic carbon (TOC), 79.53% total dissolved nitrogen (TN), 93.38% ammonia nitrogen (NH3-N) and 71.33% total dissolved phosphorus (TP) in brewery wastewater were rapidly removed by co-cultivation of T. aequale and autochthonous bacteria. Treated wastewater met the national wastewater discharge quality, and resulting algal biomass contained large amounts of high-value products chrysolaminarin, palmitoleic acid (PLA) and eicosapentaenoic acid (EPA). It is anticipated that reduced cost of algal harvesting coupled with value-added biomass could make T. aequale as a promising candidate for brewery wastewater treatment and resource utilization.

The Neuroprotective Role of Fisetin in Different Neurological Diseases: a Systematic Review.

Neurological diseases place a substantial burden on public health and have a serious impact on the quality of life of patients. Despite the multifaceted pathological process involved in the occurrence and development of these neurological diseases, each disease has its own unique pathological characteristics and underlying molecular mechanisms which trigger their onset. Thus, it is unlikely to achieve effective treatment of neurological diseases by means of a single approach. To this end, we reason that it is pivotal to seek an efficient strategy that implements multitherapeutic targeting and addresses the multifaceted pathological process to overcome the complex issues related to neural dysfunction. In recent years, natural medicinal plant-derived monomers have received extensive attention as new neuroprotective agents for treatment of neurological disorders. Fisetin, a flavonoid, has emerged as a novel potential molecule that enhances neural protection and reverses cognitive abnormalities. The neuroprotective effects of fisetin are attributed to its multifaceted biological activity and multiple therapeutic mechanisms associated with different neurological disorders. In this review article, we summarize recent research progression regarding the pharmacological effects of fisetin in treating several neurological diseases and the potential mechanisms.

The CsHSFA-CsJAZ6 module-mediated high temperature regulates flavonoid metabolism in Camellia sinensis.

High temperatures (HTs) seriously affect the yield and quality of tea. Catechins, derived from the flavonoid pathway, are characteristic compounds that contribute to the flavour of tea leaves. In this study, we first showed that the flavonoid content of tea leaves was significantly reduced under HT conditions via metabolic profiles; and then demonstrated that two transcription factors, CsHSFA1b and CsHSFA2 were activated by HT and negatively regulate flavonoid biosynthesis during HT treatment. Jasmonate (JA), a defensive hormone, plays a key role in plant adaption to environmental stress. However, little has been reported on its involvement in HT response in tea. Herein, we demonstrated that CsHSFA1b and CsHSFA2 activate CsJAZ6 expression through directly binding to heat shock elements in its promoter, and thereby repress the JA pathway. Most secondary metabolites are regulated by JA, including catechin in tea. Our study reported that CsJAZ6 directly interacts with CsEGL3 and CsTTG1 and thereby reduces catechin accumulation. From this, we proposed a CsHSFA-CsJAZ6-mediated HT regulation model of catechin biosynthesis. We also determined that negative regulation of the JA pathway by CsHSFAs and its homologues is conserved in Arabidopsis. These findings broaden the applicability of the regulation of JAZ by HSF transcription factors and further suggest the JA pathway as a valuable candidate for HT-resistant breeding and cultivation.

Metabolomics Reveals Nutritional Diversity among Six Coarse Cereals and Antioxidant Activity Analysis of Grain Sorghum and Sweet Sorghum.

Coarse cereals are rich in dietary fiber, B vitamins, minerals, secondary metabolites, and other bioactive components, which exert numerous health benefits. To better understand the diversity of metabolites in different coarse cereals, we performed widely targeted metabolic profiling analyses of six popular coarse cereals, millet, coix, buckwheat, quinoa, oat, and grain sorghum, of which 768 metabolites are identified. Moreover, quinoa and buckwheat showed significantly different metabolomic profiles compared with other coarse cereals. Analysis of the accumulation patterns of common nutritional metabolites among six coarse cereals, we found that the accumulation of carbohydrates follows a conserved pattern in the six coarse cereals, while those of amino acids, vitamins, flavonoids, and lipids were complementary. Furthermore, the species-specific metabolites in each coarse cereal were identified, and the neighbor-joining tree for the six coarse cereals was constructed based on the metabolome data. Since sorghum contains more species-specific metabolites and occupies a unique position on the neighbor-joining tree, the metabolite differences between grain sorghum 654 and sweet sorghum LTR108 were finally compared specifically, revealing that LTR108 contained more flavonoids and had higher antioxidant activity than 654. Our work supports an overview understanding of nutrient value in different coarse cereals, which provides the metabolomic evidence for the healthy diet. Additionally, the superior antioxidant activity of sweet sorghum provides clues for its targeted uses.

Unsupervised convolutional variational autoencoder deep embedding clustering for Raman spectra.

Unsupervised deep learning methods place increased emphasis on the process of cluster analysis of unknown samples without requiring sample labels. Clustering algorithms based on deep embedding networks have been recently developed and are widely used in data mining, speech processing and image recognition, but barely any of them have been used on spectra data. This study presents an unsupervised clustering algorithm for Raman spectra, called the convolutional variational autoencoder deep embedding clustering method (CVDE). It improves the network structure of the multi-layer perception (MLP) that is commonly used in other methods based on the VAE-GMM model, like VaDE, by replacing the hidden fully connected layer in the MLP with three convolution layers and two pooling layers for better clustering on the Raman spectra. The three convolution layers extend vertical channels to learn features, while pooling layers directly reduce the horizontal coding dimensions to prevent gradient explosion and overfitting. Furthermore, such network structures can easily incorporate the gradient-weighted class activation mapping (Grad-Cam) method to visualise the importance of spectral features for clustering, facilitating network tuning and spectral difference analysis. Moreover, through comparative experiments, CVDE has proven that it affords better clustering performance than current advanced clustering methods on not only the MNIST dataset but also two sets of Raman spectra: soybean oil Raman spectra with very small Raman feature differences and drug Raman spectra with a small data size. The clustering accuracies of these three datasets reach 94.48%, 90.43% and 98.70% respectively. Thus, CVDE is suitable for applications in static spectra, such as Raman spectra and LIBS spectra, and is more versatile than supervised methods in the spectral and chemical analysis fields.

Bioelectrochemical degradation of petroleum hydrocarbons: A critical review and future perspectives.

As typical pollutants, petroleum hydrocarbons that are widely present in various environmental media such as soil, water, sediments, and air, seriously endanger living organisms and human health. In the meantime, as a green environmental technology that integrates pollutant removal and resource recovery, bioelectrochemical systems (BESs) have been extensively applied to the removal of petroleum hydrocarbons from the environment. This review introduces working principles of BESs, following which it discusses the different reactor structures, application progresses, and key optimization factors when treating water, sewage sludges, sediments, and soil. Furthermore, bibliometrics was first used in this field to analyze the evolution of knowledge structure and forecast future hot topics. The research focus has shifted from the early generation of bioelectric energy to exploring mechanisms of soil remediation and microbial metabolisms, which will be closely integrated in the future. Finally, the future prospects of this field are proposed. This review focuses on the research status of bioelectrochemical degradation of petroleum hydrocarbons and provides a scientific reference for subsequent research.

OsPHR2 modulates phosphate starvation-induced OsMYC2 signalling and resistance to Xanthomonas oryzae pv. oryzae.

Phosphate (Pi) and MYC2-mediated jasmonate (JA) pathway play critical roles in plant growth and development. In particular, crosstalk between JA and Pi starvation signalling has been reported to mediate insect herbivory resistance in dicot plants. However, its roles and mechanism in monocot-bacterial defense systems remain obscure. Here, we report that Pi starvation in rice activates the OsMYC2 signalling and enhances resistance to Xanthomonas oryzae pv. oryzae (Xoo) infection. The direct regulation of OsPHR2 on the OsMYC2 promoter was confirmed by yeast one-hybrid, electrophoretic mobility shift, dual-luciferase and chromatin immunoprecipitation assays. Molecular analyses and infection studies using OsPHR2-Ov1 and phr2 mutants further demonstrated that OsPHR2 enhances antibacterial resistance via transcriptional regulation of OsMYC2 expression, indicating a positive role of OsPHR2-OsMYC2 crosstalk in modulating the OsMYC2 signalling and Xoo infection. Genetic analysis and infection assays using myc2 mutants revealed that Pi starvation-induced OsMYC2 signalling activation and consequent Xoo resistance depends on the regulation of OsMYC2. Together, these results reveal a clear interlink between Pi starvation- and OsMYC2- signalling in monocot plants, and provide new insight into how plants balance growth and defence by integrating nutrient deficiency and phytohormone signalling. We highlighted a molecular link connecting OsMYC2-mediated JA pathway and phosphate starvation signalling in monocot plant. We demonstrated that phosphate starvation promoted OsMYC2 signalling to enhance rice defence to bacterial blight via transcriptional regulation of OsPHR2 on OsMYC2.

Structural and functional organization of the MYC transcriptional factors in Camellia sinensis.

MAIN CONCLUSION: Genome-wide identification, expression analysis of the MYC family in Camellia sinensis, and potential functional characterization of CsMYC2.1 have laid a solid foundation for further research on CsMYC2.1 in jasmonate (JA)-mediated response. Myelocytomatosis (MYC) of basic helix-loop-helix (bHLH) plays a major role in JA-mediated plant growth and developmental processes through specifically binding to the G-box in the promoters of their target genes. In Camellia sinensis, studies on the MYC gene family are limited. Here, we identified 14 C. sinensis MYC (CsMYC) genes, and further analyzed the evolutionary relationship, gene structure, and motif pattern among them. The expression patterns of these CsMYC genes in different tissues suggested their important roles in diverse function in tea plant. Four MYC transcription factors with the highest homology to MYC2 in Arabidopsis were localized in the nucleus. Two of them, named CsMYC2.1 and CsMYC2.2, exhibited transcriptional self-activating activity, and, therefore, could significantly activate the promoter containing G-box motif, whereas CsJAM1.1 and CsJAM1.2 lack the transcriptional self-activating activity, indirectly mediating the JA pathway through interacting with CsMYC2.1 and CsMYC2.2. Furthermore, Yeast Two-Hybrid (Y2H) and Bimolecular Fluorescent Complimentary (BiFC) assays showed that CsMYC2.1 could interact with CsJAZ3/7/8 proteins. Genetically, the complementation of CsMYC2.1 in myc2 mutants conferred the ability to restore the sensitivity to JA signals. The results provide a comprehensive characterization of the 14 CsMYCs in C. sinensis, establishing a solid foundation for further research on CsMYCs in JA-mediated response.

SPX4 interacts with both PHR1 and PAP1 to regulate critical steps in phosphorus-status-dependent anthocyanin biosynthesis.

Phosphate (Pi) is the plant-accessible form of phosphorus, and its insufficiency limits plant growth. The over-accumulation of anthocyanins in plants is often an indication of Pi starvation. However, whether the two pathways are directly linked and which components are involved in this process await identification. Here, we demonstrate that SPX4, a conserved regulator of the Pi response, transduces the Pi starvation signal to anthocyanin biosynthesis in Arabidopsis. When phr1spx4 plants were grown under low Pi conditions, DFR expression and anthocyanin biosynthesis were induced, which distinguished the plant from the behavior reported in the phr1 mutant. We also provide evidence that SPX4 interacts with PAP1, an MYB transcription factor that controls the anthocyanin biosynthetic pathway, in an inositol polyphosphate-dependent manner. Through a physical interaction, SPX4 prevented PAP1 from binding to its target gene promoter; by contrast, during Pi-deficient conditions, in the absence of inositol polyphosphates, PAP1 was released from SPX to activate anthocyanin biosynthesis. Our results reveal a direct link between Pi deficiency and flavonoid metabolism. This new regulatory module, at least partially independent from PHR1, may contribute to developing a strategy for plants to adapt to Pi starvation.

FLOURY ENDOSPERM19 encoding a class I glutamine amidotransferase affects grain quality in rice.

As a staple food for more than half of the world's population, the importance of rice is self-evident. Compared with ordinary rice, rice cultivars with superior eating quality and appearance quality are more popular with consumers due to their unique taste and ornamental value, even if their price is much higher. Appearance quality and CEQ (cooking and eating quality) are two very important aspects in the evaluation of rice quality. Here, we performed a genome-wide association study on floury endosperm in a diverse panel of 533 cultivated rice accessions. We identified a batch of potential floury genes and prioritize one (LOC_Os03g48060) for functional analyses. Two floury outer endosperm mutants (flo19-1 and flo19-2) were generated through editing LOC_Os03g48060 (named as FLO19 in this study), which encodes a class I glutamine amidotransferase. The different performances of the two mutants in various storage substances directly led to completely different changes in CEQ. The mutation of FLO19 gene caused the damage of carbon and nitrogen metabolism in rice, which affected the normal growth and development of rice, including decreased plant height and yield loss by decreased grain filling rate. Through haplotype analysis, we identified a haplotype of FLO19 that can improve both CEQ and appearance quality of rice, Hap2, which provides a selection target for rice quality improvement, especially for high-yield indica rice varieties. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-021-01226-z.

Transcriptomic Analysis Reveals the Molecular Adaptation of Three Major Secondary Metabolic Pathways to Multiple Macronutrient Starvation in Tea (Camellia sinensis).

Tea (Camellia sinensis (L.) O. Kuntze) is a widely consumed beverage. Lack of macronutrients is a major cause of tea yield and quality losses. Though the effects of macronutrient starvation on tea metabolism have been studied, little is known about their molecular mechanisms. Hence, we investigated changes in the gene expression of tea plants under nitrogen (N), phosphate (P), and potassium (K) deficient conditions by RNA-sequencing. A total of 9103 differentially expressed genes (DEG) were identified. Function enrichment analysis showed that many biological processes and pathways were common to N, P, and K starvation. In particular, cis-element analysis of promoter of genes uncovered that members of the WRKY, MYB, bHLH, NF-Y, NAC, Trihelix, and GATA families were more likely to regulate genes involved in catechins, L-theanine, and caffeine biosynthetic pathways. Our results provide a comprehensive insight into the mechanisms of responses to N, P, and K starvation, and a global basis for the improvement of tea quality and molecular breeding.

Structural and functional insights into the LBD family involved in abiotic stress and flavonoid synthases in Camellia sinensis.

Lateral organ boundaries domain (LBD) proteins are plant-specific transcription factors that play a crucial role in growth and development, as well as metabolic processes. However, knowledge of the function of LBD proteins in Camellia sinensis is limited, and no systematic investigations of the LBD family have been reported. In this study, we identified 54 LBD genes in Camellia sinensis. The expression patterns of CsLBDs in different tissues and their transcription responses to exogenous hormones and abiotic stress were determined by RNA-seq, which showed that CsLBDs may have diverse functions. Analysis of the structural gene promoters revealed that the promoters of CsC4H, CsDFR and CsUGT84A, the structural genes involved in flavonoid biosynthesis, contained LBD recognition binding sites. The integrative analysis of CsLBD expression levels and metabolite accumulation also suggested that CsLBDs are involved in the regulation of flavonoid synthesis. Among them, CsLOB_3, CsLBD36_2 and CsLBD41_2, localized in the nucleus, were selected for functional characterization. Yeast two-hybrid assays revealed that CsLBD36_2 and CsLBD41_2 have self-activation activities, and CsLOB_3 and CsLBD36_2 can directly bind to the cis-element and significantly increase the activity of the CsC4H, CsDFR and CsUGT84A promoter. Our results present a comprehensive characterization of the 54 CsLBDs in Camellia sinensis and provide new insight into the important role that CsLBDs play in abiotic and flavonoid biosynthesis.

Kuijieling-Containing Serum Regulates Th17 and Treg Cell Differentiation by Inhibiting STAT3 Signaling In Vitro.

OBJECT: To investigate the effect of Kuijieling (KJL) on the balance between T helper 17 (Th17) and regulatory T (Treg) cells in peripheral blood mononuclear cells (PBMC) in vitro and explore the underlying mechanism. MATERIALS AND METHODS: PBMCs isolated from rats were stimulated with transforming growth factor-ß, interleukin (IL)-6, and IL-23 to induce the imbalance of Th17 and Treg cells and were treated with 10, 5, or 2.5% KJL-containing serum. The proportion of Th17 or Treg cells in CD4+ T cells was analyzed by flow cytometry, the concentrations of IL-17, IL-21, and IL-10 were assayed by ELISA, mRNA expressions of retinoic acid-related orphan receptor γt (RORγt), forkhead box protein 3 (Foxp3), and signal transducer and activator of transcription 3 (STAT3) were quantified by PCR, and phosphorylated STAT3 (p-STAT3) was analyzed by flow cytometry. RESULTS: KJL-containing serum decreased the proportion of Th17 cells and increased the proportion of Treg cells in CD4+ T cells, decreased the concentration of IL-17 and IL-21, enhanced the level of IL-10 in the cell culture supernatant, promoted the expression of Foxp3, and inhibited the levels of RORγt, STAT3, and p-STAT3. CONCLUSION: KJL suppresses the STAT3 pathway to remedy the imbalance between Th17 and Treg cells.

Effects of Resveratrol on the Mechanisms of Antioxidants and Estrogen in Alzheimer's Disease.

OBJECTIVE: To observe the effects of resveratrol (Res) on the antioxidative function and estrogen level in an Alzheimer's disease (AD) mouse model. METHODS: First, we examined the effects of Res on an AD mice model. SAMP8 mice were selected as the model, and normal-aging SAMR1 mice were used as the control group. The model mice were randomly divided into three groups: a model group, high-dose Res group (40mg/kg, intraperitoneal (ip)), and low-dose Res group (20mg/kg, ip). After receiving medication for 15 days, the mice were subjected to the water maze test to assess their spatial discrimination. The spectrophotometric method was used to detect the activity of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT) as well as the malondialdehyde (MDA) content. Quantitative PCR (q-PCR) was used to detect SOD, GSH-Px, CAT, and heme oxygenase-1 (HO-1) mRNA level changes. Western blot analysis detected HO-1 and Nrf2 protein expression. Second, we researched the effect of Res on the estrogen level in the SAMP8 model mice. The model mice were randomly divided into four groups: a model group, estrogen replacement group (0.28 mg/kg, intramuscular (im), estradiol benzoate), high-dose Res group (5 mg/kg, im), and low-dose Res group (2.5 mg/kg, im). The mice were injected, once every three days, for 5 weeks. Q-PCR was used to detect brain tissue mRNA expression changes. Western blot analysis detected ERα, ERß, and ChAT protein expression. An enzyme-linked immunosorbent assay (ELISA) kit was used to detect the expression of E2 and amyloid ß protein (Aß) in brain tissue. RESULTS: Compared with the control treatment, Res could improve the spatial abilities of the mice to a certain extent and also increase the expression of SOD, GSH-Px, CAT, and HO-1 at the mRNA level (P<0.05). In addition, enhanced SOD, GSH-Px, and CAT activities and HO-1 protein levels and decreased MDA content (P<0.05) were detected in the brain tissue of the Res-treated mice. The cytoplasmic Nrf2 content in the Res-treated mice was also decreased while the nuclear Nrf2 content and the nuclear translation rate of Nrf2 were increased (P<0.05). Res could decrease the expression of ERß in the brain tissue at the mRNA and protein levels and the expression of Aß in the brain tissue at the protein level. Res could also increase the mRNA and protein expression of ERα and ChAT and the protein expression of estradiol in the brain tissue. CONCLUSION: Res can increase the antioxidant capacity of AD models through the Nrf2/HO-1 signaling pathway. In addition, Res can enhance estrogen levels in an AD model. These findings provide a new idea for the treatment of AD.

AtHB2, a class II HD-ZIP protein, negatively regulates the expression of CsANS, which encodes a key enzyme in Camellia sinensis catechin biosynthesis.

Tea (Camellia sinensis) is an important cash crop that is beneficial to human health because of its remarkable content of catechins. The biosynthesis of catechins follows the flavonoid pathway, which is highly branched. Among the enzymes involved in catechin biosynthesis, ANTHOCYANIDIN SYNTHASE (CsANS) functions at a branch point and play a critical role. Our previous work has showed that the gene encoding CsANS is regulated by light signals; however, the molecular mechanism behind remains unclear. Here, we cloned a full-length CsANS promoter and found that it contained a cis-element recognized by Arabidopsis thaliana HOMEOBOX2 (AtHB2). AtHB2 constitutes one of the class II HOMEODOMAIN-LEUCINE ZIPPER (HD-ZIP) proteins, which accumulate in the dark and mediate the shade avoidance response in most angiosperms. To analyze the transcription of CsANS in vivo, ß-glucuronidase and luciferase reporter genes driven by the obtained promoter were introduced into A. thaliana and Nicotiana attenuata, respectively. In both expression systems there were indications that the A. thaliana PRODUCTION OF ANTHOCYANIN PIGMENT1 (AtPAP1), a MYB transcription factor of flavonoid biosynthesis, increased the activity of the CsANS promoter, while AtHB2 could significantly undermine the effect of AtPAP1. Yeast two-hybrid and bimolecular fluorescence complementation assays showed that AtHB2 interacted with the A. thaliana TRANSPARENT TESTA GLABRA 1 (AtTTG1). A yeast three-hybrid assay further suggested that AtHB2 represses the expression of CsANS and regulates its response to light signals through competitive interactions with AtTTG1. These results show that HD-ZIP II proteins participate in light regulation of flavonoid biosynthesis.

Arabidopsis PLC2 is involved in auxin-modulated reproductive development.

Phospholipase C (PLC) is an enzyme that plays crucial roles in various signal transduction pathways in mammalian cells. However, the role of PLC in plant development is poorly understood. Here we report involvement of PLC2 in auxin-mediated reproductive development in Arabidopsis. Disruption of PLC2 led to sterility, indicating a significant role for PLC2 in reproductive development. Development of both male and female gametophytes was severely perturbed in plc2 mutants. Moreover, elevated auxin levels were observed in plc2 floral tissues, suggesting that the infertility of plc2 plants may be associated with increased auxin concentrations in the reproductive organs. We show that expression levels of the auxin reporters DR5:GUS and DR5:GFP were elevated in plc2 anthers and ovules. In addition, we found that expression of the auxin biosynthetic YUCCA genes was increased in plc2 plants. We conclude that PLC2 is involved in auxin biosynthesis and signaling, thus modulating development of both male and female gametophytes in Arabidopsis.

Characterization of P2X7R and its function in the macrophages of ayu, Plecoglossus altivelis.

P2X purinoceptor 7 (P2X7R), an ATP-gated ion channel, plays an important role during the innate immune response in mammals. However, relatively little is known about the role of P2X7R in the fish immune system. Here, we cloned a cDNA sequence encoding ayu (Plecoglossus altivelis) P2X7R (aP2X7R). The predicted protein was composed of 574 amino acid residues with a P2X family signature, two transmembrane domains, and a long C-terminal. aP2X7R transcripts were mainly distributed in ayu immune tissues and significantly increased in all tested tissues and in macrophages after Listonella anguillarum infection. The aP2X7R protein was upregulated significantly in macrophages upon bacterial challenge. An antibody against the ectodomain of aP2X7R (aEPAb) and an antagonist (oATP) were used to block aP2X7R. aP2X7R siRNA was also used to knockdown the receptor expression in ayu macrophages. Cell death induced by ATP was significantly inhibited in ayu macrophages after aEPAb, oATP, or siRNA treatment. Moreover, aP2X7R ablation also resulted in suppression of phagocytic activity and ATP-induced bacterial killing in ayu macrophages. Our results indicated that aP2X7R was upregulated after infection and mediated cell death, phagocytosis, and bacterial killing of ayu macrophages.

Exine dehiscing induces rape microspore polarity, which results in different daughter cell fate and fixes the apical-basal axis of the embryo.

The roles of cell polarity and the first asymmetric cell division during early embryogenesis in apical-basal cell fate determination remain unclear. Previously, a novel Brassica napus microspore embryogenesis system was established, by which rape exine-dehisced microspores were induced by physical stress. Unlike traditional microspore culture, cell polarity and subsequent asymmetric division appeared in the exine-dehisced microspore, which finally developed into a typical embryo with a suspensor. Further studies indicated that polarity is critical for apical-basal cell fate determination and suspensor formation. However, the pattern of the first division was not only determined by cell polarity but was also regulated by the position of the ruptured exine. The first division could be equal or unequal, with its orientation essentially perpendicular to the polar axis. In both types of cell division, the two daughter cells could have different cell fates and give rise to an embryo with a suspensor, similar to zygotic apical-basal cell differentiation. The alignment of the two daughter cells is consistent with the orientation of the apical-basal axis of future embryonic development. Thus, the results revealed that exine dehiscing induces rape microspore polarization, and this polarity results in a different cell fate and fixes the apical-basal axis of embryogenesis, but is uncoupled from cell asymmetric division. The present study demonstrated the relationships among cell polarity, asymmetric cell division, and cell fate determination in early embryogenesis.

Oil body biogenesis during Brassica napus embryogenesis.

Although the oil body is known to be an important membrane enclosed compartment for oil storage in seeds, we have little understanding about its biogenesis during embryogenesis. In the present study we investigated the oil body emergence and variations in Brassica napus cv. Topas. The results demonstrate that the oil bodies could be detected already at the heart stage, at the same time as the embryos began to turn green, and the starch grains accumulated in the chloroplast stroma. In comparison, we have studied the development of oil bodies between Arabidopsis thaliana wild type (Col) and the low-seed-oil mutant wrinkled1-3. We observed that the oil body development in the embryos of Col is similar to that of B. napus cv. Topas, and that the size of the oil bodies was obviously smaller in the embryos of wrinkled1-3. Our results suggest that the oil body biogenesis might be coupled with the embryo chloroplast.