Elucidating the clinical and immunological value of m6A regulatormediated methylation modification patterns in adrenocortical carcinoma

N6-methyladenosine methylation(m6A)is a common type of epigenetic alteration that prominently affects the prognosis of tumor patients.However,it is unknown how the m6A regulator affects the tumor microenvironment(TME)cell infiltration in adrenocortical carcinoma(ACC)and how it affects the prognosis of ACC patients yet.The m6A alteration patterns of 112 ACC patients were evaluated,furthermore,the association with immune infiltration cell features was investigated.The unsupervised clustering method was applied to typify the m6A alteration patterns of ACC patients.The principal component analysis(PCA)technique was taken to create the m6A score to assess the alteration pattern in specific malignancies.We found two independent patterns of m6A alteration in ACC patients.The TME cell infiltration features were significantly in accordance with phenotypes of tumor immune-inflamed and immune desert in both patterns.The m6Ascore also served as an independent predictive factor in ACC patients.The somatic copy number variation(CNV)and patients prognosis can be predicted by m6A alteration patterns.Moreover,the ACC patients with high m6A scores had better overall survival(OS)and higher efficiency in immune checkpoint blockade therapy.Our work demonstrated the significance of m6A alteration to the ACC patients immunotherapy.The individual m6A alteration patterns analysis might contribute to ACC patients prognosis prediction and immunotherapy choice.

Potassium-Induced Regulation of Cellular Antioxidant Defense and Improvement of Physiological Processes in Wheat under Water Deficit Condition

Drought is the most common form of abiotic stress that reduces plant growth and productivity.It causes plant injuries through elevated production of reactive oxygen species(ROS).Potassium(K)is a vital plant nutrient that notably ameliorates the detrimental effect of drought stress in the plant.A pot experiment was conducted at the Laboratory of Plant Stress Responses,Faculty of Agriculture,Kagawa University,Japan,under controlled environment of green house to explore the role of K in mitigating drought severity in wheat(Triticum asevitum L.)seedlings.Three days after germination,seedlings were exposed to three water regimes viz.,100,50,and 20%field capacity(FC)for 21 days.Potassium was adjusted in Hoagland nutrient solution at 0,6 and 12 mM concentration and applied to pot instead of normal water.Results show that,water deficit stress notably reduced plant growth,biomass accumulation,leaf relative water content(RWC)along with reduced photosynthetic pigments.Increased amount of biochemical stress markers viz.,malondialdehyde(MDA),hydrogen peroxide(H_(2)O_(2)),methylglyoxal(MG),proline(Pro)as well as an impaired antioxidant defense system were observed in drought affected wheat plants.On the contrary,K supplementation resulted in improvement of biochemical and physiological parameters that worked behind in improving growth and development of the wheat plants.In addition,enzymes of ascorbateglutathione(AsA-GSH)cycle were also enhanced by supplemented K that accelerated the ROS detoxification process in plant.Although glyoxalse system did not performed well till MG was detoxified might following another short stepped pathways.Our results revealed that drought stressed plants showed better performances in terms of biochemical and physiological attributes,antioxidant defense and glyoxalase system,as well as ROS detoxification due to K supplementation with better performance at 12 mM K added in 50%FC growing condition.

Three sensitive and reliable serological assays for detection of potato virus A in potato plants

Vegetative propagation of seed potato often allows passaging of viruses to seed tubers,resulting in significant yield losses and reduction of potato tuber quality.Thus,virus detection approach is crucial for effective virus management programs and the production of virus-free seed potatoes.Among the reported potato-infecting viruses,potato virus A(PVA)is considered as one of the most important viruses in potato-growing regions worldwide.This study prepared four hybridoma lines secreting PVA-specific monoclonal antibodies(MAbs)(2D4,8E11,14A6 and 16H10)using purified PVA virions as an immunogen.Western blotting results indicated that all the four MAbs reacted strongly and specifically with the putative capsid protein of PVA.Using these four MAbs,this study developed antigen-coated plate enzyme-linked immunosorbent assay(ACP-ELISA),Dot-ELISA and Tissue print-ELISA for detection of PVA infection in potato plants.The results indicated that PVA can be detected in crude tissue extracts from infected potato plants diluted up to 1:327680(w/v,g m L^(-1))by ACP-ELISA or up to1:10240 by Dot-ELISA.The Tissue print-ELISA is the quickest and easiest approach among the three serological assays,and is more suitable for onsite large-scale potato screening programs.Further analyses of field-collected potato samples showed that the sensitivities and specificities of the three serological approaches were similar to those of RT-PCR in PVA detection and confirmed that PVA is currently widespread in Yunnan and Zhejiang provinces of China.Hence,the results strongly suggest that these highly sensitive serological approaches based on PVA-specific MAbs are useful and powerful for PVA-free seed potato production programs and PVA field surveys.

Study on Infection Ability of Rhizobium radiobacter from Rosa chinensis to Peach Tree

[Objectives] This study was conducted to analyze the infection ability of Rhizobium radiobacter from Rosa chinensis to peach tree,so as to provide a theoretical basis for the identification and control of crown gall of peach. [Methods]With peach variety ＂ Chiyohime＂ as an experimental material,R. radiobacter GOU1 was extracted from crown gall tissue of R. chinensis susceptible to crown gall,and its infection ability to peach tree was studied by in-vitro dip method.[Results]Stem segments of peach tree were inoculated with R. radiobacter GOU1 by in-vitro dip method. Calli grew rapidly after inoculation,and 28 d later,the infection rate of R. radiobacter GOU1 was 50%,indicating that R. radiobacter GOU1 could infect peach tree easier. The infection on stem segments of peach tree could be detected in the fourth week by in-vitro dip method. Culturing peach stem segments in MS medium containing 10 μM/L NAA and 10 μM/L BAP in vitro is beneficial to survival and could promote growth of callus. [Conclusions]The detection rate of R. radiobacter GOU1 infection on stem segments of peach tree did not change from the fourth week to the sixth week. Therefore,in future studies,the detection of R. radiobacter infection on inoculated stem segments of peach tree could be performed after 4 weeks of culture rather than 6 weeks of culture.

An Anionic Polymer Incorporating Low Amounts of Hydrophobic Residues Is a Multifunctional Surfactant. Part 1: Emulsifying, Thickening,Moisture-Absorption and Moisture-Retention Abilities of a FattyAcid-Containing Anionic Polysaccharide

We have been studying the function and structure of fatty acid-containing extracellular polysaccharides (FACEPS) produced by bacteria belonging to the genus Rhodococcus. In this study, we examined the relationships between the structure and emulsifying, thickening, moisture-absorption, and moisture-retention capabilities of rhodococcal FACEPS using S-2 EPS produced by R. rhodochrous strain S-2. We prepared chemically deacylated S-2 EPS (DeAcyl S-2 EPS) and palmitoylated DeAcyl S-2 EPS (ReAcyl S-2 EPS), and compared them with native S-2 EPS. All of the properties were attenuated and recovered by deacylation and reacylation of S-2 EPS, respectively. These results suggest that the fatty acid moiety of rhodococcal FACEPS is involved in such functional properties. We also showed that palmitoylation improved the emulsifying, moisture-ab-sorption, and moisture-retention abilities of other acidic polysaccharides that are commercially available. These results suggest that the acidity of the polysaccharide backbone is at least partly responsible for the observed functionality of fatty acid-containing polysaccharides. To our knowledge, this is the first report on multifunctional property of an anionic polymer incorporating low amounts of hydrophobic residues. The present findings could be useful for the creation of new multifunctional surfactants from renewable raw materials for use in various industries, e.g., in cosmetics.

An Anionic Polymer Incorporating Low Amounts of Hydrophobic Residues Is a Multifunctional Surfactant. Part 2: Emulsification, Moisture Absorption, and Moisture Retention of Alkyl Esterified Poly-γ-Glutamic Acid

In the accompanying paper (Part 1), we showed that the fatty acid moiety and the acidity of the polysaccharide-backbone of a fatty acid-containing polysaccharide might be involved in its emul-sification, moisture absorption, and moisture retention abilities. In this study, we synthesized alkyl esterified poly-γ-glutamic acid (PGA) with various chain lengths and degrees of substitution of the alkyl moieties to examine how hydrophobic groups incorporated in the anionic polymer contribute to enhanced emulsification, moisture absorption, and moisture retention. With a low degree of alkylation of PGA, these abilities were drastically improved. To improve the moisture absorption of PGA, alkylation with a short chain length is effective in forming interspaces between PGA chains to trap water molecules. Hydrophobic-hydrophilic balance may also be important to improve the emulsification and moisture retention abilities of PGA alkylates. To the best of our knowledge, this is the first report of the relationship between the structure and the multifunctional abilities of an anionic polymer incorporated with a small amount of hydrophobic residue. PGA alkylates, as well as fatty acid-containing polysaccharides, have potential use as multifunctional surfactants throughout various industries.

A Micro Peristaltic Pump Using an Optically Controllable Bioactuator

Peristalsis is widely seen in nature, as this pumping action is important in digestive systems for conveying sustenance to every corner of the body. In this paper, we propose a muscle-powered tubular micro pump that provides peristaltic transport. We utilized Drosophila melanogaster larvae that express channelrhodopsin-2 (ChR2) on the cell membrane of skeletal muscles to obtain light-responsive muscle tissues. The larvae were forced to contract with blue light stimulation. While changing the speed of the propagating light stimulation, we observed displacement on the surface of the contractile muscle tissues. We obtained peristaltic pumps from the larvae by dissecting them into tubular structures. The average inner diameter of the tubular structures was about 400 lm and the average outer diameter was about 750 lm. Contractions of this tubular structure could be controlled with the same blue light stimulation. To make the inner flow visible, we placed microbeads into the peristaltic pump, and thus determined that the pump could transport microbeads at a speed of 120 lm-s1.

Targeting Glycinebetaine for Abiotic Stress Tolerance in Crop Plants:Physiological Mechanism,Molecular Interaction and Signaling

In the era of climate change,abiotic stresses(e.g.,salinity,drought,extreme temperature,flooding,metal/metalloid(s),UV radiation,ozone,etc.)are considered as one of the most complex environmental constraints that restricts crop production worldwide.Introduction of stress-tolerant crop cultivars is the most auspicious way of surviving this constraint,and to produce these types of tolerant crops.Several bioengineering mechanisms involved in stress signaling are being adopted in this regard.One example of this kind of manipulation is the osmotic adjustment.The quarternary ammonium compound glycinebetaine(GB),also originally referred to as betaine is a methylated glycine derivative.Among the betaines,GB is the most abundant one in plants,which is mostly produced in response to dehydration caused by different abiotic stresses like drought,salinity,and extreme temperature.Glycinebetaine helps in decreased accumulation and detoxification of ROS,thereby restoring photosynthesis and reducing oxidative stress.It takes part in stabilizing membranes and macromolecules.It is also involved in the stabilization and protection of photosynthetic components,such as ribulose-1,5-bisphosphate carboxylase/oxygenase,photosystem II and quarternary enzyme and protein complex structures under environmental stresses.Glycinebetaine was found to perform in chaperone-induced protein disaggregation.In addition,GB can confer stress tolerance in very low concentrations,and it acts in activating defense responsive genes with stress protection.Recently,field application of GB has also shown protective effects against environmental adversities increasing crop yield and quality.In this review,we will focus on the role of GB in conferring abiotic stress tolerance and the possible ways to engineer GB biosynthesis in plants.

Monoclonal Antibody-Based Serological Detection of Rice Stripe Mosaic Virus Infection in Rice Plants or Leafhoppers

Rice stripe mosaic virus(RSMV) is a rhabdovirus recently found in southern part of China and can cause severe reduction in rice production. To establish serological methods for RSMV epidemiological studies and to establish a control strategy for this virus, we first purified RSMV virions from infected rice plants and then used them as an immunogen to produce four RSMV-specific monoclonal antibodies(MAbs)(i.e.,1D4, 4A8, 8E4 and 11F11). With these MAbs, we have developed a highly specific and sensitive antigen-coated plate enzyme-linked immunosorbent assay(ACP-ELISA), a Dot-ELISA and a Tissue print-ELISA for rapid detections of RSMV infection in rice plants or in leafhoppers. Our results showed that RSMV can be readily detected in RSMV-infected rice plant tissue crude extracts diluted at 1:20,971,520(w/v, g/m L)through ACP-ELISA or diluted at 1:327,680(w/v, g/m L) through Dot-ELISA. Both ACP-ELISA and Dot-ELISA can also be used to detect RSMV infection in individual RSMV viruliferous leafhopper(Recilia dorsalis) homogenate diluted at 1:307,200 and 1:163,840(individual leafhopper/l L), respectively. Detection of RSMV infection in field-collected rice samples or in RSMV viruliferous leafhoppers indicated that the three serological methods can produce same results with that produced by RT-PCR(19 of the 33 rice samples and 5 of the 16 leafhoppers were RSMV-positive). We consider that the four MAbs produced in this study are very specific and sensitive, and the three new serological methods are very useful for detections of RSMV infection in rice plants or in leafhoppers and the establishment of the disease control strategies.

Radical Cation Diels-Alder Reactions of Non-Conjugated Alkenes as Dienophiles by Electrocatalysis

Radical cation Diels-Alder reactions provide a powerful method for the construction of six-membered ring systems between both electron-rich dienes and dienophiles. However, the most recent examples of this class have been limited to β-methylstyrenes as dienophiles;the use of non-conjugated alkenes remains challenging. The present study describes the serendipitous development of novel radical cation Diels-Alder reactions by electrocatalysis that use non-conjugated alkenes as dienophiles. The key to successful transformation involves highly substituted cyclohexenyl radical cations that are stable enough to be reduced by intermolecular single electron transfer.

Systemic RNAi of V-ATPase subunit B causes molting defect and developmental abnormalities in Periplaneta fuliginosa

The vacuolar (H+)-ATPases (V-ATPases) are ATP-driven proton pumps with multiple functions in many organisms. In this study, we performed structural and functional analysis of vha55 gene that encodes V-ATPase subunit B in the smokybrown cockroach Periplaneta fuliginosa (Blattodea). We observed a high homology score of the deduced amino acid sequences between 10 species in seven orders. RNAi of the vha55 gene in R fuliginosa caused nymphal/nymphal molting defects with incomplete shedding of old cuticles, growth inhibition, as well as bent and wrinkled cuticles of thoraxes and abdominal segments. Since growth inhibition caused by vha55 RNAi did not interfere in the commencement of cockroach molting, molting timing and body growth might be controlled by independent mechanism. Our study suggested V-ATPases might be a good candidate molecule for evolutionary and developmental studies of insect molting.

The Calcineurin B-Like Calcium Sensors CBL1 and CBL9 Together with Their Interacting Protein Kinase ClPK26 Regulate the Arabidopsis NADPH Oxidase RBOHF

Stimulus-specific accumulation of second messengers like reactive oxygen species （ROS） and Ca^＋ are central to many signaling and regulation processes in plants. However, mechanisms that govern the reciprocal interrelation of Ca^＋ and ROS signaling are only beginning to emerge. NADPH oxidases of the respiratory burst oxidase homolog （RBOH） family are critical components contributing to the generation of ROS while Calcineurin B-like （CBL） Ca^＋ sensor proteins together with their interacting kinases （CIPKs） have been shown to function in many Ca^＋- signaling processes. In this study, we identify direct functional interactions between both signaling systems. We report that the CBL-interacting pro- tein kinase ClPK26 specifically interacts with the N-terminal domain of RBOHF in yeast two-hybrid analyses and with the full-length RBOHF protein in plant cells. In addition, CIPK26 phosphorylates RBOHF in vitro and co-expression of either CBL1 or CBL9 with CIPK26 strongly enhances ROS production by RBOHF in HEK293T cells. Together, these findings identify a direct interconnection between CBL-ClPK-mediated Ca^＋ signaling and ROS signaling in plants and provide evidence for a synergistic activation of the NADPH oxidase RBOHF by direct Ca^＋-binding to its EF-hands and Ca2＋-induced phospho-rylation by CBL1/9-ClPK26 complexes.

凤果花叶病毒高灵敏度血清学检测技术

目的:建立基于单克隆抗体的检测番茄等植物中凤果花叶病毒(PepMV)的血清学方法,为PepMV的田间调查和诊断及其科学防控提供快速实用的检测技术。创新点:首次制备了抗PepMV的高度特异和灵敏的单克隆抗体,并利用制备的单抗建立了3种能特异且灵敏地检测PepMV的血清学方法。方法:以差速离心方法提纯的PepMV粒子作为免疫原免疫BALB/c小鼠,通过杂交瘤技术获得了能稳定传代并分泌PepMV单克隆抗体的杂交瘤细胞株;杂交瘤细胞注射到小鼠腹腔获得单克隆抗体腹水,并以制备单抗为核心,根据血清学原理建立检测植物中PepMV的双抗夹心酶联免疫吸附试验(DAS-ELISA)、斑点酶联免疫吸附试验(Dot-ELISA)和组织印迹酶联免疫吸附试验(Tissue print-ELISA)三种血清学检测方法;利用田间番茄样品分析建立的血清学方法检测PepMV的有效性。结论:利用杂交瘤技术获得了6株能分泌高度特异灵敏PepMV单克隆抗体的杂交瘤细胞株,以分泌的单抗为核心建立了检测植株中PepMV的DASELISA,Dot-ELISA和Tissue print-ELISA三种高度灵敏的血清学新技术。三种建立的血清学技术检测感染PepMV的番茄植株均呈强阳性反应,而检测健康番茄及感染其他5种植物病毒的植株呈阴性反应,且DAS-ELISA和Dot-ELISA血清学技术检测番茄病叶粗提液的灵敏度分别达到1:1310720和1:20480倍稀释(质量体积比,g/mL)。田间样品检测结果发现,建立的血清学技术的检测结果与反转录聚合酶链反应(RT-PCR)的检测结果一致,表明建立的血清学方法可有效地用于植物中PepMV的检测。同时,本研究首次发现PepMV己在我国云南番茄作物上发生流行。PepMV单克隆抗体的制备及其灵敏血清学检测方法的建立有益于PepMV的田间调查和诊断及其科学防控。

A Novel Thermomorphic System for Electrocatalytic Diels-Alder Reactions

The discovery that lithium bis(trifluoromethane)sulfonamide (LiTFSI)/1-nitropropane (PrNO2) solution functions as a less polar alternative to lithium perchlorate (LiClO4)/nitromethane (MeNO2) solution has led to the development of a novel thermomorphic system for electrocatalytic Diels-Alder reactions. Methyl cyclohexane (Me-c-Hex) can form a monophasic condition with LiTFSI/PrNO2 solution at room temperature, enabling the use of hydrophobic dienophiles. After the electrochemical reaction, a biphasic condition can be formed at –50℃, where the cycloadducts are selectively recovered from the upper Me-c-Hex phase and the remaining lower LiTFSI/PrNO2 solution can be reused.