Wickerhamomyces anomalus has garnered significant interest for its remarkable ability to shape wine flavor profiles. However, during fermentation, yeast cells are inevitably exposed to ethanol stress. Nitric oxide (NO) is a key signaling molecule that plays diverse physiological roles in living organisms. However, its impact on cell wall and membrane homeostasis under ethanol stress remains unclear. In this study, we investigated the regulatory effects of NO on the cellular integrity of W. anomalus under ethanol stress by supplementing with the NO donor (SNP) or the NO scavenger (Carboxy-PTIO) using physiological analyses focused on the cell wall and membrane. Our results demonstrated that elevated NO levels alleviated ethanol-induced morphological and ultrastructural alterations, primarily by maintaining cellular homeostasis, ultimately promoting cell survival and vitality. Specifically, NO mitigated ethanol stress-compromised cell wall integrity by activating the cell wall integrity (CWI) pathway and increasing intracellular levels of β-glucan and chitin. Furthermore, NO alleviated ethanol-induced disruption of membrane homeostasis by remodeling membrane composition, enhancing integrity, permeability, and fluidity, while simultaneously increasing ATPase activity, elevating intracellular K⁺ levels, and regulating fatty acid synthesis. These findings provide crucial insights into the mechanistic basis of NO-regulated stress responses in W. anomalus under ethanol stress and offer a foundation for developing novel strategies to improve its industrial utilization under fermentative stress conditions.

Wickerhamomyces anomalus, a flavor-modulating non-Saccharomyces yeast, has garnered significant interest for its remarkable ability to shape wine flavor profiles. However, during fermentation, yeast cells are inevitably exposed to ethanol stress. The specific structural consequences of this stress, particularly its impact on cell wall and membrane homeostasis, remain unclear. In this study, we investigated the effects of ethanol stress on the cellular integrity of W. anomalus through a physiological analysis focused on the cell wall and membrane. Our results demonstrated that ethanol stress induced significant morphological and ultrastructural alterations, which were primarily attributed to the disruption of cellular homeostasis. Specifically, ethanol stress compromised cell wall integrity, activated the cell wall integrity (CWI) pathway, and increased the intracellular levels of β-glucan and chitin. Furthermore, ethanol stress disrupted membrane homeostasis by remodeling its composition, reducing integrity and fluidity, while increasing permeability and simultaneously enhancing ATPase activity and elevated intracellular K⁺ levels. Fatty acid profiling also demonstrated a decrease in the monounsaturated fatty acid C18:1 and an increase in very long-chain fatty acids (VLCFAs; C22:0 and C24:0) under ethanol challenge. Exogenous supplementation of these fatty acids was shown to enhance the ethanol stress tolerance of W. anomalus. These findings provide crucial insights into the mechanistic basis of ethanol stress response in W. anomalus and offer a foundation for developing novel strategies to improve its industrial utilization under fermentative stress conditions.

Wickerhamomyces anomalus, a flavor-modulating non-Saccharomyces yeast used in winemaking, experiences escalating ethanol stress during fermentation, yet the physiological and structural consequences remain unclear. This study assessed responses to 9% (v/v) ethanol stress with and without exogenous antioxidants (10 mM N-acetylcysteine and 2.5 mM glutathione) by measuring: reactive oxygen species (ROS) via fluorescence staining; superoxide dismutase (SOD) and catalase (CAT) activities plus glutathione content colorimetrically; mitochondrial membrane potential using rhodamine 123; ultrastructure via transmission electron microscopy; electron transport chain complex activities and ATP levels colorimetrically; and autophagy via monodansylcadaverine staining. Results indicated ethanol exposure induced ROS overproduction and oxidative stress while concurrently activating SOD and catalase activities and increasing GSH. Furthermore, ethanol reduced mitochondrial membrane potential, inhibited electron transport chain activity, decreased ATP synthesis, and triggered autophagy. Conversely, antioxidant supplementation alleviated oxidative damage, restored mitochondrial function, suppressed autophagy, and enhanced cell viability. These findings elucidate ethanol's mechanistic toxicity in yeast and provide a foundation for developing antioxidant-enhanced, ethanol-tolerant W. anomalus strains for industrial fermentation.

Tyrosine (Tyr) degradation is a crucial pathway in animals. However, its role in plants remains to be examined. Fumarylacetoacetate hydrolase (FAH) is the final enzyme involved in Tyr degradation. Studies of a mutant of the SHORT-DAY SENSITIVE CELL DEATH 1 (SSCD1) gene encoding FAH in Arabidopsis have shown that blockage of this pathway results in the accumulation of Tyr metabolites, thereby inducing cell death under short-day conditions. Seed dormancy is a critical trait which is regulated by endogenous and environmental cues, among which abscisic acid (ABA) and gibberellin (GA) are the primary effectors. ABA induces seed dormancy, whereas GA releases seed dormancy. In this study, sscd1 seeds displayed deep dormancy and hypersensitivity to the GA biosynthesis inhibitor paclobutrazol, but not to ABA during germination. However, exogenous GA3 could not completely recover dormancy or germination of sscd1 seeds. Moreover, GA3 level was reduced, which was consistent with the decreased expression of GA3-oxidase 1 in imbibed sscd1 seeds. Furthermore, SSCD1 acted upstream of RGA-LIKE 2. Eliminating the accumulation of Tyr metabolites by inhibiting homogentisate dioxygenase, an enzyme upstream of FAH, completely rescued the phenotype of sscd1 seeds. Additionally, germination of sscd1 seeds was hypersensitive to FAH deficiency-induced accumulation of succinylacetone, which is a Tyr metabolite. These findings suggest that FAH deficiency in sscd1 causes accumulation of Tyr metabolites, thereby disrupting GA biosynthesis and signaling. This resulted in deep dormancy and hypersensitivity to paclobutrazol during germination and highlights the important role of the Tyr degradation pathway in GA-mediated seed dormancy and germination.

Sesquiterpene lactone artemisinin is one of the main agents used to treat malaria. Artemisinin is produced in the glandular secretory trichomes (GSTs) of Artemisia annua. The plant hormones and metabolic pathways regulate the artemisinin content of A. annua. It was possible to examine the functions of auxin, an important plant hormone, in the development of GSTs in A. annua by enhancing the expression of iaaM, which encodes a tryptophan monooxygenase involved in the biosynthesis of auxin. Additionally, the effects of co-overexpression of aldehyde dehydrogenase 1 (ALDH1), P450 monooxygenase (CYP71AV1), and iaaM on the density, length, and width of GSTs and the contents of artemisinin were further investigated. Results indicated that overexpression of iaaM might increase the density, length, and width of GSTs by enhancing auxin biosynthesis. This study also proved the key regulatory role of ALDH1 in the biosynthesis of artemisinin. Moreover, co-overexpression of ALDH1, CYP71AV1, and iaaM successfully increased the density, length, and width of GSTs and improved the artemisinin content in A. annua. Therefore, we established a theoretical basis for modifying artemisinin accumulation in this study by regulating the expression of auxin and artemisinin synthesis-related genes using a metabolic engineering method.

Complement factor I (CFI), a complement inhibitor, is well known for regulating the complement system activation by degrading complement component 3b (C3b) in animal serum, thus becoming involved in innate defense. Nevertheless, the functional mechanisms of CFI in the complement system and in host-pathogen interactions are far from being clarified in teleost fish. In the present study, we cloned and characterized the CFI gene, CiCFI, from grass carp (Ctenopharyngodon idella) and analyzed its function in degrading serum C3b and expression changes after grass carp reovirus (GCRV) infection. The open reading frame of CiCFI was found to be 2121 bp, encoding 706 amino acids with a molecular mass of 79.06 kDa. The pairwise alignments showed that CiCFI shared the highest identity (66.9%) with CFI from Carassius gibelio and the highest similarity (78.7%) with CFI from Danio rerio. The CiCFI protein was characterized by a conserved functional core Tryp_SPc domain with the catalytic triad and substrate binding sites. Phylogenetic analysis indicated that CiCFI and the homologs CFIs from other teleost fish formed a distinct evolutionary branch. Similar with the CFIs reported in mammals, the recombinant CiCFI protein could significantly reduce the C3b content in the serum, demonstrating the conserved function of CiCFI in the complement system in the grass carp. CiCFI mRNA and protein showed the highest expression level in the liver. After GCRV infection, the mRNA expressions of CiCFI were first down-regulated, then up-regulated, and then down-regulated to the initial level, while the protein expression levels maintained an overall downward trend to the late stage of infection in the liver of grass carps. Unexpectedly, the protein levels of CiCFI were also continuously down-regulated in the serum of grass carps during GCRV infection, while the content of serum C3b proteins first increases and then returns to the initial level, suggesting a distinct role of CiCFI in regulating complement activation and fish-virus interaction. Combining our previous results that complement factor D, a complement enhancer, shows continuously up-regulated expression levels in grass carps during GCRV infection, and this study may provide the further essential data for the full picture of complex complement regulation mechanism mediated by Df and CFI of the grass carp during pathogen infection.

Nectar phenolics have a widespread effect on honey bees and their colonies. Because of their complex, non-linear interactions, it is difficult to assess honey bee health risks from exposure to real-world floral nectar with complex phenolic mixture. In the study, we investigate the bee losses of Apis mellifera in the flowering period of the Mexican sunflower Tithonia diversifolia in southwestern China, and use data mining approach to model the relationships between nectar phenolics and bee losses. The results show that bee losses are closely related to the phenolics of isochlorogenic acid, p-coumaric acid, chlorogenic acid and galangin, identified from the sunflower nectar. The nectar phenolics do not cause bee-poisoning to death, but can trigger bee colonies to explore food sources at risk. Also, each of these phenolics acts in a dichotomous mode, with above a certain value destructing colonies and below such value affecting little. This study provides new insight into the mechanism underlying the catastrophic events of bee losses or honey harvests, which have been reported worldwide.

Abstract Honeybees play a significant role in the plant–pollinator interactions of many flowering plants. The ecological and evolutionary consequences of plant–pollinator interactions vary by geographic region, and the effects of honeybees on the reproduction of toxic plants have not been well studied. We measured the florescence of toxic plants, the flower-visiting behaviour of honeybees and the effects of pollination on the fertility, weight and moisture content of seeds. The effects of climatic factors on the number of flowers, and the spatial and temporal variation in pollinator visits were evaluated, and the effects of pollinator visits on seed quality were evaluated. Flower visitors were diverse, climatic factors had a great impact on spatio-temporal flowering variation and the number of bee visits was strongly correlated with the spatio-temporal variation in the number of flowers. Honeybees strongly increase the fullness and weight of seeds. Our study demonstrated a good ecological fit between the spatio-temporal variation in the flowering of toxic plants and the general validity of honeybee pollination syndrome in the south of Hengduan Mountains in East Asia. A linear relationship between honeybee visitation and plant reproduction can benefit the stabilization of plant reproduction.

电子类专业的毕业实习是教学中重要环节,专业对口的实习可以提升学生的专业技能,提高学生就业的竞争力.鉴于目前的整体形势,社会对电子类专业人才需求大,但提供实习岗位不多.实习的对口性对学校和学生来说都是迫切需要解决的问题.提出了利用校内资源,建设电子类毕业实习平台的方案,用以缓解该难题.该方案经过了实践检验,取得了初步的成效,可为同类院校电子类专业毕业实习提供一定的参考.

Leaves and flowers of Amorphophallus konjac do not develop simultaneously thus unique features can be elucidated through study of flowering transformation in A. konjac. In this study, transcriptome libraries of A. konjac leaf buds (LB) and flower buds (FB) were constructed followed by high-throughput sequencing. A total of 68,906 unigenes with an average length of 920 bp were obtained after library assembly. Out of these genes, 24,622 unigenes had annotation information. A total of 6859 differentially expressed genes (DEGs) were identified through differential expression analysis using LB as control. Notably, 2415 DEGs were upregulated whereas 4444 DEGs were downregulated in the two transcriptomes. Go and KEGG analysis showed that the DEGs belonged to 44 functional categories and were implicated in 98 metabolic pathways and 38 DEGs involved in plant hormone signal transduction. Several genes were mined that may be involved in A. konjac flower bud differentiation and flower organ development. Eight DEGs were selected for verification of RNA-seq results using qRT-PCR analysis. Two FLOWERING LOCUS T ( FT ) genes named AkFT1 and AkFT2 were identified though homologous analysis may be the florigen gene implicated in modulation of A. konjac flowering. These genes were significantly upregulated in flower buds compared with the expression levels on leaf buds. Overexpression of AkFT genes though heterologous expression in Arabidopsis showed that the transgenics flowered at a very early stage relative to wild type plants. These findings indicate that AkFT1 and AkFT2 function as regulation genes in A. konjac flowering development and the two genes may present similar functions during flowering transition.

α-Galactosidase (EC 3.2.1.22) refers to a group of enzymes that hydrolyze oligosaccharides containing α-galactoside-banded glycosides, such as stachyose, raffinose, and verbascose. These enzymes also possess great potential for application in sugar production, and in the feed and pharmaceutical industries. In this study, a strain of Lactosphaera pasteurii (WHPC005) that produces α-galactosidase was identified from the soil of Western Hunan, China. It was determined that the optimal temperature and pH for this α-galactosidase were 45 °C and 5.5, respectively. The activity of α-galactosidase was inhibited by K+, Al3+, Fe3+, fructose, sucrose, lactose, galactose, SDS, EDTA, NaCl, and (NH4)2SO4, and enhanced by Ca2+, Fe2+, Mn2, Zn2+, glucose, and raffinose. The optimal inducer was raffinose, and the optimal induction concentration was 30 μmol/L. The α-galactosidase gene was cloned using random fragment cloning methods. Sequence analysis demonstrated that the open reading frame of the α-galactosidase gene was 1230 bp, which encodes a putative protein of 409 amino acids in length. Bioinformatics analysis showed that the isoelectric point and molecular weight of this α-galactosidase were 4.84 and 47.40 kD, respectively. Random coils, alpha helixes, and beta turns were observed in its secondary structure, and conserved regions were found in the tertiary structure of this α-galactosidase. Therefore, this α-galactosidase-producing bacterial strain has the potential for application in the feed industry.

将Ca2+响应实时荧光报告系统引入雌二醇诱导生长素结合蛋白(ABP1)调控表达的BY2细胞中,获得了 ABP1过表达(ABP1-ox)、抑制表达(ABP1-anti)同时对Ca2+标记的几个BY2细胞株.对这些细胞株进行雌二醇诱导调控其ABP1过表达或抑制表达后,分析ABP1的表达量,并通过在细胞外添加IAA处理,实时观察ABP1调控表达后细胞的Ca2+信号变化.结果表明:ABP1-ox细胞在雌二醇诱导后细胞内ABP1表达量显著提高,细胞经IAA处理后,细胞膜内、核膜周围均有迅速而强烈的荧光信号,说明ABP1过表达后细胞能快速响应胞外的IAA作用,将信号转导到细胞内,使细胞质内Ca2+信号迅速增强;而ABP1-anti细胞经雌二醇诱导后,细胞内ABP1表达显著下调,细胞经IAA处理后,相对于对照,细胞内的荧光信号微弱且呈点状,细胞核附近的荧光不明显,说明细胞内Ca2+信号的转导受到抑制.这证明BY2细胞中ABP1参与生长素信号与细胞内Ca2+响应的信号转导过程.

为研究青蒿素合成的关键酶基因AaADS过表达对黄花蒿腺毛发育和青蒿素含量的影响,本研究根据GenBank收录的黄花蒿AaADS基因启动子和cDNA序列设计引物,采用同源克隆的方法从黄花蒿品种(428-A)中扩增出AaADS基因启动子和cDNA序列.构建GUS报告载体AaADSpro∷GUS和植物表达载体AoADSpro∷AaADS,利用农杆菌介导的叶盘转化法转化黄花蒿.通过对转基因植株叶片腺毛的GUS基因活性检测和不同组织器官AaADS基因RT-PCR半定量分析,以及过表达AaADS基因的植株叶片下表皮腺毛荧光显微观察、统计和植株青蒿素含量的测定,结果表明:AaADS基因能在黄花蒿腺毛细胞中高效表达;转基因植株中AaADS基因表达量明显高于对照,且在叶片组织表达量最高;转基因植株叶片腺毛细胞较对照植株明显增大,过表达AaADS基因植株中青蒿素含量达27.3 mg/g DW,比对照植株中青蒿素含量12 mg/gDW提高了 2.3倍.本研究说明AaADS基因过表达能有效促进黄花蒿腺毛细胞的生长和青蒿素的生物合成,为选育高青蒿素含量的黄花蒿新品系提供可行性方法.

酪氨酸降解途径在动物生长发育中发挥着重要作用.然而,该途径在植物中的功能还需进一步鉴定.马来酰乙酰乙酸异构酶是酪氨酸降解途径中的关键酶.ζ类谷胱甘肽S-转移酶(GSTZ)具有马来酰乙酰乙酸异构酶活性.为探讨酪氨酸降解途径在植物响应盐胁迫中的作用,本研究分析了拟南芥(Arabidopsis thaliana) AtGSTZ1基因在盐胁迫下的表达特征以及AtGSTZ1过表达植株对盐胁迫的耐受性,发现:AtGSTZ1表达受盐胁迫诱导;在盐胁迫下,AtGSTZ1过表达植株具有较高的种子萌发率和子叶绿化率;此外,盐胁迫下过表达植株积累了较少的H2O2,且过氧化氢酶基因CAT1表达量和过氧化氢酶活性都高于野生型.实验结果表明,AtGSTZ1通过调控盐胁迫下H2O2的积累而影响拟南芥的耐盐性.本研究结果为证明酪氨酸降解途径在植物响应盐胁迫中的作用提供依据.

The greater wax moth, Galleria mellonella (Linnaeus, 1758), is a notorious pest of honey bee colonies that has negatively affected the global apicultural industry.Olfactory cues influence the behavior of wax moth, where males attract females, making them an ideal candidate for pheromone studies.However, the molecular mechanism of chemoreception in G. mellonella pertaining to sex pheromone recognition has not been elucidated.In this study, transcriptome sequencing was conducted on the antennae of male and female greater wax moths to assess the differential expression patterns of chemosensory genes and better understand the underlying olfactory mechanism.In the results, a total of 121 chemosensory gene transcripts were identified, including 37 odorant-binding proteins, 35 chemosensory proteins, 33 olfactory receptors, 14 ionotropic receptors and 2 sensory neuron membrane proteins.The expression patterns of these genes were determined using the estimated fragments per kilobase of transcript per million fragments mapped.Among the 114 DEGs, 66 were expressed exclusively in the female antennae, whereas the remaining were expressed predominantly in the male antennae.Additionally, five chemosensory-related genes (OBP69alike, OBP72-like, CSP7, CSP10 and OR29) were differentially expressed between the two samples.In conclusion, the study lay a foundation for understanding the olfactory functions of chemosensory genes in G. mellonella, which can help to control and prevent the damage caused by this pest.

The plant pathogenesis-related (PR) proteins play a crucial role in the defense of plants against pathogens and orchestrate the innate immune system of plants. In this paper, a non-normalized cDNA library of the leaf was constructed to obtain a comprehensive view of PR proteins of Macleaya cordata. Specifically, 511 expressed sequence tags (ESTs) were generated using Sanger sequencing. All ESTs were assembled into 364 non-redundancy sequences, including 78 clusters and 286 singlets. The PR protein expression profile of the medicinal herb M. cordata has been investigated and is represented by defensin, lipid-transfer protein, (S)-norcoclaurine synthase, and major allergen protein, suggesting that the herb contains rich active proteins against pathogens. Furthermore, two defensins were selected for recombinant expression in yeast, and the antimicrobial activities were explored. Since they both present a broad antimicrobial spectrum, they are of particular importance for agricultural and medicinal applications. Our study describes defensins in Papaveraceae for the first time and provides novel insights into the effective components. In addition to the alkaloids, PR proteins (such as defensins, lipid transfer proteins, (S) - norcoclaurine synthase, major allergen protein, and Class IV chitinases) are involved in the antibacterial and anti-inflammatory activities of M. cordata.

Heteropoda venatoria in the family Sparassidae is highly valued in pantropical countries because the species feed on domestic insect pests. Unlike most other species of Araneomorphae, H. venatoria uses the great speed and strong chelicerae (mouthparts) with toxin glands to capture the insects instead of its web. Therefore, H. venatoria provides unique opportunities for venom evolution research. The venom of H. venatoria was explored by matrix-assisted laser desorption/ionization tandem time-of-flight and analyzing expressed sequence tags. The 154 sequences coding cysteine-rich peptides (CRPs) revealed 24 families based on the phylogenetic analyses of precursors and cysteine frameworks in the putative mature regions. Intriguingly, four kinds of motifs are first described in spider venom. Furthermore, combining the diverse CRPs of H. venatoria with previous spider venom peptidomics data, the structures of precursors and the patterns of cysteine frameworks were analyzed. This work revealed the dynamic evolutionary trends of venom CRPs in H. venatoria: the precursor has evolved an extended mature peptide with more cysteines, and a diminished or even vanished propeptides between the signal and mature peptides; and the CRPs evolved by multiple duplications of an ancestral ICK gene as well as recruitments of non-toxin genes.

为研究如何降低风力机叶片表面风压与噪声,应用数值模拟方法研究在风力机叶片前缘表面施加等离子体激励对风力机叶片表面风压与噪声的抑制作用.结果表明:叶片的噪声主要产生在叶尖压力面前缘负压区.如施加前缘射向后缘方向等离子体激励,叶片压力面前缘负压峰值及中部负压略微增大,噪声声功率级增大;而施加后缘射向前缘方向等离子体激励,叶片压力面前缘负压峰值及中部负压减小,噪声声功率级减小.来流风速为10 m/s,风力机叶片在17.5 r/min转速下施加前缘射向后缘方向等离子体激励后风力机叶片噪声最多可降低7.72 dB(12.19％),在14 r/min转速条件下施加后缘射向前缘方向等离子体激励后风力机叶片噪声最多可降低7.56 dB(15.25％).

1 前言 不同群势蜂群产子育虫数量是蜂群生物学中的一项重要研究内容,根据不同群势蜂群的繁殖特点,采取不同的管理方法和生产策略,对蜂群增殖,夺取蜂蜜高产,具有重要的指导作用. 关于不同群势蜂群的产子育虫数量,前苏联学者在西方蜜蜂上做了一系列研究,指出: “虽然蜂群内育虫数量随蜂群群势的增长,其总量是增加的,但其生长速度却逐渐放缓” “按蜜蜂单位计算,较小的蜂群比较大的蜂群培育蜂子数量较多,生长得较快.”

Extracellular protease Vpr (Vpr), gamma-glutamyltranspeptidase (GGT; EC 2.3.2.2) and glyoxal/methylglyoxal reductase (YvgN; EC 1.1.1.21) are extracellular enzymes involved in feather degradation, which were identified by secretome analyses from an efficient feather-degrading strain Bacillus subtilis CH-1. The encoding sequences corresponding to the three secretory enzymes were cloned into vector pET22b for recombinant expression in Escherichia coli strain BL21 (DE3). Afterward, the proteins containing the C-terminal His-tag were purified using a Ni-IDA column. The optimal temperatures and pH values for protease activity of recombinant Vpr, GGT, and YvgN were identified as 45 °C/pH 7.0, 40 °C/pH 8.0, and 50 °C/pH 6.0 respectively when casein is the substrate. Furthermore, the synergistic effects of the three enzymes were studied using feather powder as substrate. Vpr was the core enzyme to hydrolyze keratin, while GGT and YvgN were coenzymes providing reducing activities for keratin decomposition. The keratinolytic activity was enhanced to about 1.4-folds when YvgN and Vpr applied together in comparison to Vpr alone. And the keratinolytic activity almost reached to 1.5-folds when all the three enzymes were combined to use. The study provides a novel perspective of the mechanism of keratin degradation by microorganisms, and thereby may also be of relevance for the design of an industrial process for enzymatic keratin degradation; however, additional experiments must be done to substantiate this conclusion.