Melatonin may increase disease resistance and flavonoid biosynthesis through effects on DNA methylation and gene expression in grape berries.

BACKGROUND: Melatonin can regulate plant growth, development and biotic responses by causing global changes in gene expression; however, the melatonin-induced changes in gene expression via the modification of DNA methylation remain unclear in plants. RESULTS: A total of 1,169,852 and 1,008,894 methylated cytosines (mCs) were identified in the control and melatonin-treated grape berries, respectively, and mCs occurred primarily at CG sites, followed by CHG sites and CHH sites. Compared to the control, melatonin treatment broadly decreased methylation levels at CHG and particularly CHH sites in various gene regions. Melatonin treatment generated a total of 25,125 differentially methylated regions (DMRs), which included 6517 DMR-associated genes. RNA-Seq demonstrated that 2479 genes were upregulated, and 1072 genes were repressed by melatonin treatment. The evaluation of the interconnection of the DNA methylome and transcriptome identified 144 genes showing a negative correlation between promoter methylation and gene expression, which were primarily related to biotic stress responses and flavonoid biosynthesis. Additionally, the application of 5́-azacytidine and melatonin led to similar effects on mycelial growth of B. cinerea, berry decay rate and flavonoid biosynthesis. Moreover, EDS1 was used to show that melatonin increased gene expression by decreasing promoter methylation levels. CONCLUSION: Our results demonstrated that melatonin broadly decreased DNA methylation and altered gene expression in grape berries. We propose that melatonin increases disease resistance and flavonoid biosynthesis by decreasing the methylation levels of the promoters of the genes involved.

Growth mechanism and photoelectric properties of a silver nanowire network prepared by solid state ionics method.

A macroscopic silver nanowire (AgNW) network is grown by solid state ionics method. The ion flow during growth of the AgNW network is controlled by maintaining a current in the order of 10-7 A. Scanning electron microscopy (SEM) analysis reveals that the growth direction of AgNWs in the network is irregular and spread evenly in all directions and the nanowires are 40-160 nm in diameter. The microcosmic mechanism of silver nanostructures grown by the solid state ionics method is established by real time and in situ SEM analysis of the growth process of the AgNW networks. To study the photoelectric properties of the network, a self-supported AgNW network sample (∼1 mm wide and 8 mm long) is irradiated with lasers of different wavelengths of 375, 405, 532, 633, 808, and 1064 nm and 10.6 µm, and changes in the current between the two ends of the sample are recorded. The network displays negative photoconductance effect, and the maximum light responsivity is 43 mA W-1. The network displays light responsivity in the ultraviolet light-to-mid-infrared light region, with response times of tens of milliseconds. These findings indicate that the AgNW network has broad application prospect in ultra-wide spectrum photoelectric detection.

Facile fabrication of eutectic gallium-indium alloy nanostructure and application in photodetection.

Eutectic gallium-indium (EGaIn) alloy is a kind of liquid metal and has attracted much attention due to good properties. In order to satisfy the trend of miniaturization and realize more practical applications, the exploration for preparation method and properties of EGaIn at nanoscale are very important. Here, facile vacuum thermal evaporation method is developed to fabricate EGaIn nanostructures. The EGaIn nanoparticle and nanofilm with naturally formed 5 nm thick oxide layers are well prepared. The oxide film formed on the EGaIn surface is an important factor, making the properties of the nanostructure different from the properties of the bulk. Compared with ignorance of oxide layer in bulk materials, the proportion of oxide layer increases evidently in nanostructures, which produce obvious influence on the electric and optical properties. The rectifying characteristic and optoelectronic performance are experimentally observed. The EGaIn nanostructures can generate evident photocurrent responses with good responsivities (∼1 mA W-1) and response speed (∼1 s) under irradiation of 206 nm, 405 nm, 532 nm, 635 nm, 808 nm, 1064 nm and 10.6 µm lasers. These properties are completely different from the metallic properties of EGaIn bulk material.

Transcriptomic and phosphoproteomic profiling and metabolite analyses reveal the mechanism of NaHCO3-induced organic acid secretion in grapevine roots.

BACKGROUND: Organic acid secretion is a widespread physiological response of plants to alkalinity. However, the characteristics and underlying mechanism of the alkali-induced secretion of organic acids are poorly understood. RESULTS: Oxalate was the main organic acid synthesized and secreted in grapevine (a hybrid of Vitis amurensis, V. berlandieri and V. riparia) roots, while acetate synthesis and malate secretion were also promoted under NaHCO3 stress. NaHCO3 stress enhanced the H+ efflux rate of grapevine roots, which is related to the plasma membrane H+-ATPase activity. Transcriptomic profiling revealed that carbohydrate metabolism was the most significantly altered biological process under NaHCO3 stress; a total of seven genes related to organic acid metabolism were significantly altered, including two phosphoenolpyruvate carboxylases and phosphoenolpyruvate carboxylase kinases. Additionally, the expression levels of five ATP-binding cassette transporters, particularly ATP-binding cassette B19, and two Al-activated malate transporter 2 s were substantially upregulated by NaHCO3 stress. Phosphoproteomic profiling demonstrated that the altered phosphoproteins were primarily related to binding, catalytic activity and transporter activity in the context of their molecular functions. The phosphorylation levels of phosphoenolpyruvate carboxylase 3, two plasma membrane H+-ATPases 4 and ATP-binding cassette B19 and pleiotropic drug resistance 12 were significantly increased. Additionally, the inhibition of ethylene synthesis and perception completely blocked NaHCO3-induced organic acid secretion, while the inhibition of indoleacetic acid synthesis reduced NaHCO3-induced organic acid secretion. CONCLUSIONS: Our results demonstrated that oxalate was the main organic acid produced under alkali stress and revealed the necessity of ethylene in mediating organic acid secretion. Additionally, we further identified several candidate genes and phosphoproteins responsible for organic acid metabolism and secretion.

Self-assembled gold micro/nanostructure arrays based on superionic conductor RbAg4I5 films.

Herein, we propose a new strategy to fabricate gold (Au) micro/nanostructure arrays by photocatalytic solid-state electrochemical reaction between superionic conductor RbAg4I5 and Au films. The Au and RbAg4I5 films were successively deposited on a clean quartz substrate by vacuum thermal deposition method. A copper microgrid possessing periodic holes 100 µm in diameter was put above the RbAg4I5 film as a mask plate, whereupon irradiation from a 405 nm wavelength laser was used to diffuse gold ions (Au+ ions) into vacant silver sites of RbAg4I5 and transfer Au+ through ion passageways in the RbAg4I5 film. When the laser was turned off, the Au+ ions were easily reduced due to low activity compared to the silver (Ag+) ions. After multiple on/off cycles of the 405 nm laser, the irradiated area of uniform Au film exhibited a periodic structural unit array whose period was the same as that of the mask plate hole array. Atomic force microscope and scanning electron microscope images revealed that a self-assembled needle-like nanostructure array grew perpendicular to the substrate surface inside each circle's structural unit. The height of the grown nanostructure array increased with laser power density. Raman enhancement of the gold nanostructure array as substrate was detected using Rhodamine 6G (R6G) ethanol solutions as probe molecules. The enhancement effect increased with the height of the grown nanostructure array, and could increase by two orders of magnitude greater than that of unirradiated Au film. This strategy offers a new method for the micro/nanostructure processing of gold and provides microscale-array-mediated surface-enhancement Raman-scattering (SERS) substrates comprising Au nanostructures for application in high-sensitivity spectrum analysis.

[Intensity loss of two-photon excitation fluorescence microscopy images of mouse oocyte chromosomes].

As an optical microscope with high resolution, two-photon excitation (TPE) fluorescence microscope is widely used in noninvasive 3D optical imaging of biological samples. Compared with confocal laser scanning microscope, TPE fluorescence microscope provides a deeper detecting depth. In spite of that, the image quality of sample always declines as the detecting depth increases when a noninvasive 3D optical imaging of thicker samples is performed. Mouse oocytes with a large diameter, which play an important role in clinical and biological fields, have obvious absorption and scattering effects. In the present paper, we performed compensation for two-photon fluorescence images of mouse oocyte chromosomes. Using volume as a parameter, the attenuation degree of these chromosomes was also studied. The result of our data suggested that there exists a severe axial intensity loss in two-photon microscopic images of mouse oocytes due to the absorption and scattering effects. It is necessary to make compensation for these images of mouse oocyte chromosomes obtained from two-photon microscopic system. It will be specially needed in studying the quantitative three-dimensional information of mouse oocytes.

Effects of piperine and deoxyschizandrin on synchronized Ca²âº oscillations in cultured hippocampal neuronal cells.

It has been reported that piperine (PIP) and deoxyschizandrin (DS) can modulate synchronized Ca²âº oscillations in cultured hippocampal neuronal networks. We investigated the modulation effects of four different combinations of piperine and deoxyschizandrin on synchronized Ca²âº oscillations in cultured hippocampal neuronal networks. The results showed that all four combinations (PIP:DS 4.9:1.9, 2.45:2.85, 7.35:0.95, and 2.45:0.95 mg/L) inhibit Ca²âº oscillation intensity to a similar extent. However, the first three combinations had strong inhibitory effects on the frequency of Ca²âº oscillations whereas the last combination (2.45:0.95 mg/L) only slightly enhanced the frequency of Ca²âº oscillations. We propose an improved Chay's model to explain the mechanism of the effects of piperine and deoxyschizandrin on synchronized Ca²âº oscillations in cultured hippocampal neuronal cells. We concluded that deoxyschizandrin modulated synchronized Ca²âº oscillations in cultured hippocampal neuronal networks bidirectionally and the effect depended on concentration. Deoxyschizandrin reduced voltage-gated sodium channel conductance and ATP-sensitive potassium channel conductance, and affected the rate of exchange of intracellular calcium and the pump activity of Ca²âº-ATPase in the endoplasmic reticulum (ER). Piperine reduced the activity of calcium release in the ER, and reduced the pump activity of calcium in the cytomembrane or enhanced the pump activity of Ca²âº-ATPase in the ER.

[A method for real-time observation of intracellular free Ca2+ concentration in single bovine retinal neurons].

In order to observe the neurotrophin's influence on intracellular free calcium concentration, the test neuron was labeled by fluorescence indicator Fluo-3, and imaged by self-built real-time fluorescence microscopy system. The authors observed the changes in intracellular free calcium concentration ([Ca2+]i) in the bovine retinal neurons under the influence of four kinds of neurotrophins such as brain derived neurotrophic factor BDNF. On account of the fact that fluorescence indicator's intensity decays over time, it is necessary to apply a "decay removal correction" to the fluorescence intensity in order to show the fluorescence intensity that solely represents [Ca2+]i. The corrected data shows an increase after adding neurotrophins to neurons, which is consistent with similar results published by other groups. Thus, our method of imaging living cells is feasible, and "decay removal correction" is reliable.

VvMYB14 participates in melatonin-induced proanthocyanidin biosynthesis by upregulating expression of VvMYBPA1 and VvMYBPA2 in grape seeds.

This work demonstrated that melatonin increases continuously in seeds, particularly seed coats, during berry ripening. Exogenous melatonin treatments significantly increased the proanthocyanidin (PA) content, partially through ethylene signaling, in seed coats. VvMYB14 expression exhibited patterns similar to melatonin accumulation over time, which was largely induced by melatonin treatment in seed coats during berry ripening. Additionally, VvMYB14 bound to the MBS element of the VvMYBPA1 promoter to activate expression. VvMYB14 overexpression largely upregulated expression of VvMYBPA1, VvMYBPA2 and VvLAR1 and increased the PA content in grape seed-derived calli. Similar increases in AtTT2 and AtBAN expression and PA content were found in VvMYB14-overexpressing Arabidopsis seeds. It was also observed that VvMYB14 overexpression increased ethylene production and thereby induced expression of VvERF104, which bound to the ERF element of the VvMYBPA2 promoter and activated its expression. Additionally, VvERF104 suppression reduced the VvMYB14 overexpression-induced increases in expression of VvMYBPA2 and VvLAR1 and PA content. Further experiments revealed that melatonin-induced increases in the expression of VvMYBPA1, VvMYBPA2, VvERF104 and VvLAR1 and PA accumulation were significantly reduced in VvMYB14-suppressing grape calli and leaves. Collectively, VvMYB14 mediates melatonin-induced PA biosynthesis by directly transactivating VvMYBPA1 expression and indirectly upregulating VvMYBPA2 expression via VvERF104.

Effect of Petroleum Generation and Retention on Nanopore Structure Change in Laminated and Massive Shales-Insights from Hydrous Pyrolysis of Lacustrine Source Rocks from the Permian Lucaogou Formation.

The lacustrine shale of the Middle Permian Lucaogou Formation in the Jimusar Sag is the principal prospective unconventional target in the Junggar Basin. The effect of petroleum generation and retention on nanopore structure change during thermal maturity in lacustrine shale is still unclear. In this study, two laminated and two massive shale samples from the Permian Lucaogou Formation were selected to study this change by closed hydrous pyrolysis. The pyrolysis temperatures were 295, 320, 345, 370, and 400 °C, which cover from the mature to the post-mature stage. Total organic carbon (TOC), Rock-Eval, and low-pressure N2 adsorption tests on pyrolyzed shale samples before and after extractable organic matter (EOM) extraction were conducted systematically. The results indicate that (1) the petroleum generation on nanopore structure change is in stages. The peak nanopore volume expanding stage is the late oil window (R o = 0.9-1.35%). At the post-mature stage (R o > 1.35%), the mesopore volume decreased and the majority of the nanopore space is from macropores. (2) The presence of EOM decreased both mesopores and macropores in the peak oil window. (3) The organic-rich laminated shale generated more macropores than massive shale with increasing thermal maturity. The results of this study shed light on the dynamic effect of laminae fabric, petroleum generation, and retention on shale nanopore structure change across the oil window.

[Visualization of the age-related changes in expressions of DNA methyltransferases in mouse oocytes using two-photon imaging system].

Quantum dot (QD) is widely used as fluorescent labeling dye for its strong anti-fluorescence quenching, high quantum yield, wide absorption spectra, and narrow emission spectra. In the present paper, QD 585-labeled DNA methyltransferases (Dnmts) and Hoechst 33342-labeled chromosomes were imaged simultaneously using two-photon imaging system. The authors' results showed that aging mouse oocytes may be not suitable for in-vitro maturation: both the localizations and expression levels of Dnmts in in-vitro matured oocytes of aging mice were changed, and these changes may be related to the abnormal DNA methylation modification.

Research on China-EU equipment manufacturing Trade dependence in intra-industry specialization view.

As the foundation of the industrial economy, the equipment manufacturing industry takes an important position on the China-EU trade. Based on the analysis of the overall trend and structure of China-EU equipment manufacturing industry trade in 2007-2020, this article involves the construction of trade concentration into trade dependence metrics, and then calculate the degree of interdependence between China and EU equipment manufacturing trade in 2020. The perspective of the intra-industry specialization will be used to analyze China-EU equipment manufacturing trade dependency in 2020. The results show that: (1) Although China-EU equipment manufacturing trade has continued to grow, China had an imbalanced export structure to the EU, and electronic equipment exports are too high; (2) Regardless of import or export, the trade dependence of the EU countries on China about equipment manufacturing was higher than that of China on European countries; (3) China mainly depended on the EU about the high-end equipment manufacturing trade, which brings risks to Chinese manufacturing supply chains.

Local large temperature difference and ultra-wideband photothermoelectric response of the silver nanostructure film/carbon nanotube film heterostructure.

Photothermoelectric materials have important applications in many fields. Here, we joined a silver nanostructure film and a carbon nanotube film by van der Waals force to form a heterojunction, which shows excellent photothermal and photoelectric conversion properties. The local temperature difference and the output photovoltage increase rapidly when the heterojunction is irradiated by lasers with wavelengths ranging from ultraviolet to terahertz. The maximum temperature difference reaches 215.9 K, which is significantly higher than that of other photothermoelectric materials reported in the literature. The photothermal and photoelectric responsivity depend on the wavelength of lasers, which are 175~601 K W-1 and 9.35~40.4 mV W-1, respectively. We demonstrate that light absorption of the carbon nanotube is enhanced by local surface plasmons, and the output photovoltage is dominated by Seebeck effect. The proposed heterostructure can be used as high-efficiency sensitive photothermal materials or as ultra-wideband fast-response photoelectric materials.

Quantitative analysis of intracellular calcium and mitochondrial kinetic fluorescence changes in GSNO-induced thymocyte early apoptosis.

A fluorescence microscopy imaging technique was applied to observe the single-cell kinetic changes of intracellular Ca(2+) concentration ([Ca(2+)](i)) and mitochondrial membrane potential (ΔΨ(m)) during the early stage of S-nitrosoglutathione (GSNO)-induced thymocytes apoptosis. The kinetic features of [Ca(2+)](i) and ΔΨ(m) were quantitatively analyzed and compared by fitting the fluorescence intensity data. The mathematical parameter, inflection point which indicated the time point when [Ca(2+)](i) or ΔΨ(m) changed the most rapidly, was proposed to analyze the fitting curve. The results revealed that the inflection point of [Ca(2+)](i) always appeared prior to that of ΔΨ(m) during apoptosis induced by a certain GSNO concentration. Both the [Ca(2+)](i) and ΔΨ(m) changed in a GSNO concentration-dependent manner. Another parameter, half-max effect point was also employed and displayed the similar results. Such quantitative analyses of real-time observations at the single-cell level are useful for interpreting the sequence of the biological events operating in GSNO-induced thymocyte apoptosis.

Melatonin alters the secondary metabolite profile of grape berry skin by promoting VvMYB14-mediated ethylene biosynthesis.

The interplay between melatonin and ethylene in the regulation of fruit metabolism and the underlying molecular mechanism of this interplay remain largely unclear. Here, widely targeted metabolomics analysis revealed a total of 464 metabolites present in berry skin. Among them, 27 significantly differentially accumulated metabolites (DAMs) were produced in response to melatonin treatment in the presence or absence of 1-MCP. Most of the DAMs were secondary metabolites, including flavonoids, phenolic acids, stilbenes, and flavonols. Additionally, the accumulation of 25 DAMs was regulated by melatonin via ethylene. RNA-seq analysis indicated that melatonin primarily regulated the pathways of plant hormone signal transduction and secondary metabolite biosynthesis via ethylene. Gene-metabolite association analysis showed that melatonin regulated the expression of the VvSTS1, VvF3H, VvLAR2, and VvDFR genes, suggesting that these genes may play key roles in regulating secondary metabolites in the skin; additionally, VvMYB14 and VvACS1 were suggested to be involved in the regulation of secondary metabolites. Further experiments revealed that melatonin induced the expression of VvMYB14 and that VvMYB14 increased ethylene production by transcriptionally activating VvACS1, thereby affecting the accumulation of secondary metabolites. Collectively, melatonin promotes ethylene biosynthesis and alters secondary metabolite accumulation through the regulation of VvACS1 by VvMYB14.

Ultra-wideband self-powered photodetector based on suspended reduced graphene oxide with asymmetric metal contacts.

The ultraviolet to terahertz band forms the main focus of optoelectronics research, while light detection in different bands generally requires the use of different materials and processing methods. However, researchers are aiming to realize multi-band detection simultaneously in the same device in certain specific application scenarios and ultra-wideband photoelectric detectors can also realize multi-function and multi-system integration. Therefore, the research and development work on ultra-wideband photoelectric detectors has important practical application value. Here, we produced self-powered suspended Pd-reduced graphene oxide-Ti (Pd-rGO-Ti) photodetectors. We varied the properties of the rGO films by using different annealing temperatures and achieved p-doping and n-doping of the films by evaporating palladium films and titanium films, respectively, thus enabling preparation of photothermoelectric (PTE) photodetectors based on rGO films. The resulting detectors have excellent photoelectric responses over a wideband illumination wavelength range from 375 nm to 118.8 µm (2.52 THz). At the same time, we determined the best experimental conditions and device structure by varying the channel width, the laser spot irradiation position and the experimental atmospheric pressure. The maximum responsivity obtained from our detectors is 142.08 mV W-1, the response time is approximately 100-200 ms and the devices have high detection sensitivity. Based on this work, we assumed in the subsequent experiments that detectors with higher performance can be obtained by reducing the channel width and atmospheric pressure. With advantages that include simple fabrication, low cost, large-scale production potential and ultra-wideband responses, these Pd-rGO-Ti photodetectors have broad application prospects in high-performance integrated optoelectronics.

[Observation on cytoskeleton and root tip growth of Arabidopsis thaliana by two-photon laser scanning microscopy].

Cytoskeleton plays an important role in plant cells and their movements, developments, and so on. With MS culture medium, the present paper cultured GFP-fused Arabidopsis thaliana in asepsis condition, which completed its whole lifecycle in the lab, starting from germination, going through plant growth, blossom, and ending at fructification. During this process, the authors used two-photon laser scanning microscopy (TPLSM) which was suitable for 4D observation on large, thick, live samples to observe cytoskeleton in seed, root tip, vessel and root hair of the Arabidopsis thaliana. Dynamic root tip growth as well as the growing speed was also observed. Applying IAA to the Arabidopsis thaliana, the authors found that the growing speed of root tip was enhanced by about 3.8 times.

[DNA nanosensor fluorescence imaging microscopy].

Quantum dots have many excellent optical properties such as high quantum yield, long fluorescence lifetime, wide excitation spectrum and narrow emission spectrum, tunable emission wavelength and so on, thus have become a newpopular type of fluorescence probes in these years. Quantum-dot-based DNA nanosensor comprising streptavidin-conjugated quantum dots, capture probes with biotin and reporter probes with Cy5 was designed to detect DNA or RNA segments. Capture probes and reporter probes were connected by the target DNA or RNA segments so that quantum dots and Cy5s could be together and FRET (fluorescence resonance energy transfer) could be detected. In the present work, quantum-dot-based DNA nanosensor was combined with ICCD fluorescence microscopy imaging system through the authors' experiments. Using the total internal reflection fluorescence (TIRF), FRET between quantum dots and Cy5s was recorded by ICCD showing that segments of singlestranded target DNA with 30-base length were detected in solution using DNA nanosensor. When Cy5-ssDNA-Biotins were added into streptavi din-conjugated quantum dots in solution, by real time recording, the FRET efficiency was found to increase with time, which indicated the process of streptavidin-conjugated quantum dots capturing Cy5-ssDNA-Biotins. It was also observed that streptavidin-conjugated quantum dots and Cy5-ssDNA-Biotins could both enter living Chinese hamster ovary cells and have FRET. The process of streptavidin-conjugated quantum dots capturing Cy5-ssDNA-Biotins was detected in the cells as well and Cy5s were photobleached after a long time of irradiation. It has been proved that detecting DNA or RNA segments in living cells with DNA nanosensor is possible.

[Observation of double-stained African green monkey kidney COS-7 cells using total internal reflection double-channel fluorescence microscopy].

Using a Dual-View wavelength splitter, double-stained African green monkey kidney COS-7 cells, transfected with pEGFP-Myosin 15a and costained with Rhodamine-filopodia were observed based on an ICCD(intensified charge couple device) fluorescence micro-imaging systems. Total internal reflection fluorescence microscopy was used to observe the overexpression of Myosin 15a to the tips of the elongation filopodia. An approach to collecting fluorescence in two channels and avoiding spectra crosstalk was employed to observe Myosin 15a and filopodia distribution in African green monkey kidney COS-7 cells. High sensitivity TIRF technology and two channels imaging method provided a wide application in bio-medical studies.