Protoplast Membrane Research Articles

Cell integrity maintenance and genetic transfection of protoplasts in an acoustofluidic system

Hydrodynamic force loading platforms based on acoustofluidics have been developed to study the mechanical deformation of cancer cells and to control cell behavior. However, so far there have been no experimental measurements on living plant cells using such an acoustic approach. Unique structures, including cell walls, allow plant cells to exhibit more variation in mechanical resistance. In this work, we analyzed plant cell deformation and membrane permeability using a gigahertz (GHz) acoustofluidic system. By recording the proportion of intact cells in the cell population, we evaluated the mechanical resistance of the protoplasts to the hydrodynamic forces of the acoustic streaming. The results showed that a regenerated primary cell wall (PCW) could significantly improve the mechanical resistance of individual plant cells within 24 h compared to the freshly prepared protoplasts without walls. The results of enzymatic degradation showed that three main components of the primary cell wall contribute to different degrees to the improvement of the mechanical properties of the cells, in decreasing order: cellulose, hemicellulose, and pectin. Furthermore, we have shown that such an acoustofluidic system can alter the permeability of the protoplast membrane in a controllable manner for transient gene expression.

A Carbonic Anhydrase, ZmCA4, Contributes to Photosynthetic Efficiency andModulates CO2 Signaling Gene Expression by Interacting with Aquaporin ZmPIP2;6 inMaize.

Carbonic anhydrase (CA) catalyzes the reversible CO2 hydration reaction that produces bicarbonate for phosphoenolpyruvate carboxylase (PEPC). This is the initial step for transmitting the CO2 signal in C4 photosynthesis. However, it remains unknown whether the maize (Zea mays L.) CA gene, ZmCA4, plays a role in the maize photosynthesis process. In our study, we found that ZmCA4 was relatively highly expressed in leaves and localized in the chloroplast and the plasma membrane of mesophyll protoplasts. Knock-out of ZmCA4 reduced CA activity, while overexpression of ZmCA4 increased rubisco activity, as well as the quantum yield and relative electron transport rate in photosystem II. Overexpression of ZmCA4 enhanced maize yield-related traits. Moreover, ZmCA4 interacted with aquaporin ZmPIP2;6 in bimolecular fluorescence complementation and co-immunoprecipitation experiments. The double-knock-out mutant for ZmPIP2;6 and ZmCA4 genes showed reductions in its growth, CA and PEPC activities, assimilation rate and photosystem activity. RNA-Seq analysis revealed that the expression of other ZmCAs, ZmPIPs, as well as CO2 signaling pathway homologous genes, and photosynthetic-related genes was all altered in the double-knock-out mutant compared with the wild type. Altogether, our study's findings point to a critical role of ZmCA4 in determining photosynthetic capacity and modulating CO2 signaling regulation via its interaction with ZmPIP2;6, thus providing insight into the potential genetic value of ZmCA4 for maize yield improvement.

Overexpression of a C3HC4‐type E3‐ubiquitin ligase contributes to salinity tolerance by modulating Na+ homeostasis in rice

AbstractSoil salinity has a negative effect on crop yield. Therefore, plants have evolved many strategies to overcome decreases in yield under saline conditions. Among these, E3‐ubiquitin ligase regulates salt tolerance. We characterized Oryza sativa Really Interesting New Gene (RING) Finger C3HC4‐type E3 ligase (OsRFPHC‐4), which plays a positive role in improving salt tolerance. The expression of OsRFPHC‐4 was downregulated by high NaCl concentrations and induced by abscisic acid (ABA) treatment. GFP‐fused OsRFPHC‐4 was localized to the plasma membrane of rice protoplasts. OsRFPHC‐4 encodes a cellular protein with a C3HC4‐RING domain with E3 ligase activity. However, its variant OsRFPHC‐4C161A does not possess this activity. OsRFPHC‐4‐overexpressing plants showed enhanced salt tolerance due to low accumulation of Na+ in both roots and leaves, low Na+ transport in the xylem sap, high accumulation of proline and soluble sugars, high activity of reactive oxygen species (ROS) scavenging enzymes, and differential regulation of Na+/K+ transporter expression compared to wild‐type (WT) and osrfphc‐4 plants. In addition, OsRFPHC‐4‐overexpressing plants showed higher ABA sensitivity under exogenous ABA treatment than WT and osrfphc‐4 plants. Overall, these results suggest that OsRFPHC‐4 contributes to the improvement of salt tolerance and Na+/K+ homeostasis via the regulation of changes in Na+/K+ transporters.

TaNRAMP3 is essential for manganese transport in Triticum aestivum

Manganese (Mn) is an essential trace element for almost all living organisms. In plants, Mn deficiency, which is occurs in calcareous soils or alkaline soils, severely limiting crop yields. However, the potential mechanism of Mn transport in Triticum aestivum is still obscure. Here, we found that TaNRAMP3, a member of the naturally resistant macrophage protein (NRAMP) family in Triticum aestivum, is located in the plasma membrane of protoplasts and functions as an influx transporter for Mn in yeast (Δsmf1). The expression of TaNRAMP3 was induced under Mn-deficiency conditions. Furthermore, TaNRAMP3-RNAi plants exhibited a sensitive phenotype, while transgenic plants overexpressing TaNRAMP3 showed a tolerant phenotype. In addition, TaNRAMP3 rescued the sensitive phenotype of Arabidopsis nramp1 mutant under Mn deficiency condition. In summary, our study reveals the key role of TaNRAMP3 in Mn transport in Triticum aestivum, allowing it to adapt to Mn-deficiency stress. These findings provide new insights for the cultivation of Mn-deficiency tolerant wheat varieties.

Silencing of a Wheat Ortholog of Glucan Synthase-Like Gene Reduced Resistance to Blumeria graminis f. sp. tritici.

Wheat powdery mildew, caused by the obligate biotrophic ascomycete fungal pathogen Blumeria graminis f. sp. tritici (Bgt), is a major threat to wheat production worldwide. It is known that Arabidopsis thaliana glucan synthase-like 5 (AtGSL5) improves the resistance of wheat to powdery mildew by increasing its anti-penetration abilities. However, the function of glucan synthase-like (GSL) orthologs in crop species remains largely unknown. In this study, TaGSL22, a novel functional ortholog of AtGSL5, was isolated as the only Bgt-induced GSL gene in wheat. Phylogenetic analysis indicated that TaGSL22 was conserved within the group of Gramineae and showed a closer relationship to GSL orthologs from monocots than to those from dicots. The TaGSL22 transcript was highest in the wheat leaves, followed by stems then roots. TaGSL22 was localized in the cell membrane and cytoplasm of wheat protoplasts, as predicted by transmembrane structure analysis. In addition, expression of TaGSL22 was induced by the plant hormones ethylene (ETH) and salicylic acid (SA), but down-regulated by jasmonate (JA) and abscisic acid (ABA). The transcript level of TaGSL22 was up-regulated in the incompatible interaction between Bgt and wheat, whereas it remained relatively unchanged in the compatible interaction. Knocking down of TaGSL22 by virus-induced gene silencing (VIGS) induced a higher infection type in the wheat–Bgt interaction. The TaGSL22-silenced plants exhibited reduced resistance to Bgt, accompanied by decreased callose accumulation. Our study shows a conserved function of GSL genes in plant immunity associated with penetration resistance, and it indicates that TaGSL22 can be used to improve papilla composition and enhance resistance to wheat powdery mildew.

The rice phosphoinositide-specific phospholipase C3 is involved in responses to osmotic stresses via modulating ROS homeostasis

Four members of phosphoinositide-specific phospholipase C (PI-PLC) are predicted in rice genome. Although the involvement of OsPLC1 and OsPLC4 in the responses of rice to salt and drought stresses has been documented, the role of OsPLC3 in which, yet, is elusive. Here, we report that OsPLC3 was ubiquitously expressed in various tissues during the development of rice. The expression of YFP-tagged OsPLC3 was observed at the plasma membrane (PM), cytoplasm and nucleus of rice protoplasts, onion epidermal cells and tobacco leaves. The catalytic activity of OsPLC3 was measured using the thin-layer chromatography (TLC) method. The inhibition of OsPLC3 expression was detected in the treatments of NaCl and mannitol. Overexpression (OE) of OsPLC3 produced plants showing more sensitive to osmotic stresses when they were compared to the wild-type (HJ) and osplc3 mutants, the phenomena such as decreased plant fresh weight and increased water loss rate (WLR) were observed. Under the treatment of NaCl or mannitol, expressions of a subset osmotic stress-related genes were altered, in both OE and osplc3 mutant lines. In addition, the expressions and the enzyme activities of reactive oxygen species (ROS) scavengers were significantly decreased in OE lines, leading to over-accumulation of ROS together with less osmotic adjustment substances including proline, soluble sugars and soluble proteins in OE plants which caused the growth inhibition. Thus, our results suggested that, via modulating ROS homeostasis, OsPLC3 is involved in responses to the osmotic stress in rice.

A tomato receptor-like cytoplasmic kinase, SlZRK1, acts as a negative regulator in wound-induced jasmonic acid accumulation and insect resistance.

Jasmonates accumulate rapidly and act as key regulators in response to mechanical wounding, but few studies have linked receptor-like cytoplasmic kinases (RLCKs) to wound-induced jasmonic acid (JA) signaling cascades. Here, we identified a novel wounding-induced RLCK-XII-2 subfamily member (SlZRK1) in tomato (Solanum lycopersicum) that was closely related to Arabidopsis HOPZ-ETI-DEFICIENT 1 (ZED1)-related kinases 1 based on phylogenetic analysis. SlZRK1 was targeted to the plasma membrane of tobacco mesophyll protoplasts as determined by transient co-expression with the plasma membrane marker mCherry-H+-ATPase. Catalytic residue sequence analysis and an in vitro kinase assay indicated that SlZRK1 may act as a pseudokinase. To further analyse the function of SlZRK1, we developed two stable knock-out mutants by CRISPR/Cas9. Loss of SlZRK1 significantly altered the expression of genes involved in JA biosynthesis, salicylic acid biosynthesis, and ethylene response. Furthermore, after mechanical wounding treatment, slzrk1 mutants increased transcription of early wound-inducible genes involved in JA biosynthesis and signaling. In addition, JA accumulation after wounding and plant resistance to herbivorous insects also were enhanced. Our findings expand plant regulatory networks in the wound-induced JA production by adding RLCKs as a new component in the wound signal transduction pathway.

Fine mapping and candidate gene analysis of maize defective kernel mutant <italic>dek54</italic>

<p indent=0mm>Maize kernel is closely related to yield and nutritive quality. Study on the function of maize kernel development relative genes provides important basis for the molecular mechanism analysis, yield increasing and nutritive quality improving. B73 pollen was treated with ethyl methylmethanesulfonate (EMS) and a defective maize kernel <italic>defective kernel 54</italic> (<italic>dek54</italic>) was screened. <italic>dek54</italic> had small mature kernel, wrinkled and whitened seed coat phenotype. Genetic analysis indicated that <italic>dek54</italic> is a recessive mutant controlled by a single gene. Paraffin sections showed starchy endosperm cells of <italic>dek54</italic> had irregular shape and dense arrangement at developmental stage. Scanning electron microscopy observation indicated that protein bodies around starch granules in the central region of the <italic>dek54 </italic>mature kernel endosperm were fewer and arranged more loosely compare to wild type. Total protein, zein, amino acids components contents and total nitrogen content of <italic>dek54 </italic>mature kernel were significantly lowered compared with the wild type. <italic>dek54</italic> was located on chromosome 7 within the interval of the physical distance of about 290 kb between markers SSR6 and SSR7. Sequencing revealed that the 351th base G on the 2nd exon of <italic>Zm00001d019294 </italic>gene changed into A, which led to the premature termination of the protein translation. <italic>Zm00001d019294</italic> gene was specifically expressed in immature maize kernel, and has the highest expression in 12 DAP (days after pollination) immature kernel. Targeted mutation was performed using CRISPR/Cas9 system to identify that mutant phenotype was caused by candidate gene <italic>Zm00001d019294</italic>. <italic>Dek54</italic> encoded an MFS (major facilitator superfamily) protein and had high homology with ZmNRT1.5 (nitrate transporter). Besides, Dek54 protein was localized in the plasma membrane of maize protoplasts. The study of <italic>dek54 </italic>laid the foundation for the molecular mechanism analysis of maize kernel development.

OrMKK3 Influences Morphology and Grain Size in Rice.

Although morphology and grain size are important to rice growth and yield, the identity of abundant natural allelic variations that determine agronomically important differences in crops is unknown. Here, we characterized the function of mitogen-activated protein kinase 3 from Oryza officinalis Wall. ex Watt encoded by OrMKK3. Different alternative splicing variants occurred in OrMKK3. Green fluorescent protein (GFP)–OrMKK3 fusion proteins localized to the cell membrane and nuclei of rice protoplasts. Overexpression of OrMKK3 influenced the expression levels of the grain size-related genes SMG1, GW8, GL3, GW2, and DEP3. Phylogenetic analysis showed that OrMKK3 is well conserved in plants while showing large amounts of variation between indica, japonica, and wild rice. In addition, OrMKK3 slightly influenced brassinosteroid (BR) responses and the expression levels of BR-related genes. Our findings thus identify a new gene, OrMKK3, influencing morphology and grain size and that represents a possible link between mitogen-activated protein kinase and BR response pathways in grain growth.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12374-020-09290-2.

TaYS1A, a Yellow Stripe-Like Transporter Gene, Is Required for Wheat Resistance to Puccinia striiformis f. sp. Tritici.

Yellow stripe-like (YSL) transporters are required for the transportation of metal-phytosiderophores and are structurally related to metal-nicotianamine complexes. Some studies also reported the involvement of YSL transporters in pathogen-induced defense. However, the molecular mechanisms of YSL genes involved in biotic stress responses are still not clear, especially in cereal crops. This study aimed to functionally characterize TaYS1A during the interaction of wheat and Puccinia striiformis f. sp. tritici (Pst), the causal agent of stripe rust disease. TaYS1A was localized in the cell membrane of wheat protoplasts and Nicotiana benthamiana cells. TaYS1A was significantly up-regulated in wheat leaves after being infected with the avirulent Pst isolate CYR23 and after treatment with salicylic acid (SA). Silencing of TaYS1A by the virus-induced gene silencing method enhanced the susceptibility of wheat to Pst accompanied by reducing the accumulation of SA and H2O2 and down-regulating the transcriptions of TaPR1 and TaPR2. In addition, TaYS1A was found to interact with TaNH2, a homolog of OsNH2, by yeast-two-hybrid assay, and silencing of TaYS1A diminished the expression of TaNH2. Our findings suggested the existence of positive regulation of TaYS1A in providing resistance against Pst by modulating SA-induced signaling and offered new insight into the biological role of YSL in wheat against pathogens.

Functional analysis of rubber tree receptor-like cytoplasmic kinase HbBIK1 in plant root development and immune response

Nature rubber from rubber tree is one of the most important raw materials for industry. However, rubber trees have been seriously affected by fungal diseases, resulting in large yield loss of nature rubber. Therefore, it is important to identify the disease resistance–related genes for genetically engineered breeding of disease resistance in rubber tree. In this study, the homologous gene of Arabidopsis thaliana receptor-like kinase AtBIK1, HbBIK1, was selected from rubber tree. The full length of HbBIK1 gene was 1260 bp of 419 aa with an intracellular kinase motif. HbBIK1 expression was induced by Oidium heveae Steinmann infection, indicating that HbBIK1 might be involved in response to O. heveae Steinmann infection. The expression level of HbBIK1 was also induced by jasmonic acid (JA) treatment, but not by salicylic acid (SA) and ethylene (ET) treatments. In addition, HbBIK1 was specifically localized on plasma membrane of rubber tree mesophyll protoplasts. Transgenic HbBIK1 lines using AtBIK1 native promoter could rescue not only root length but also disease sensitivity to Alternaria brassicicola in A. thaliana bik1 mutant, indicating the functional similarity between AtBIK1 and HbBIK1. These results showed that HbBIK1 might involve in plant disease resistance and could be used as a candidate disease-related gene for defense genetic breeding in rubber tree.

Ca2+ efflux is negatively correlated with apple firmness

Calcium plays an important role in the regulation of fruit firmness, but the mechanism behind this remains poorly understood. In this study, we investigated the firmness of the fruit pulp in the peduncle, calyx, and equatorial parts of Fuji apples. The pulp protoplasts of the three parts were obtained by enzymatic hydrolysis. The patch clamp technique was used to measure the Ca2+ current on the protoplast membranes of the three parts. The firmness was significantly different among the three fruit parts. The equatorial part, which had the lowest firmness, had the highest Ca2+ efflux, while the lowest Ca2+ efflux was measured near the calyx, which had the highest firmness. Supplementation of potassium fertilizer to apple trees promoted fruit firmness and reduced Ca2+ efflux from pulp cells. Spraying 10 mg/L auxin directly onto young fruit increased the firmness and reduced cell Ca2+ efflux. These results indicated that Ca2+ efflux is correlated with fruit firmness. Because fruit softening is a natural phenomenon during fruit ripening, we conclude that the reduction in cellular Ca2+ efflux can maintain cell viability and increase the firmness of apple fruit pulp.

Functional Analysis of the Soybean GmCDPK3 Gene Responding to Drought and Salt Stresses.

Plants have a series of response mechanisms to adapt when they are subjected to external stress. Calcium-dependent protein kinases (CDPKs) in plants function against a variety of abiotic stresses. We screened 17 CDPKs from drought- and salt-induced soybean transcriptome sequences. The phylogenetic tree divided CDPKs of rice, Arabidopsis and soybean into five groups (I–V). Cis-acting element analysis showed that the 17 CDPKs contained some elements associated with drought and salt stresses. Quantitative real-time PCR (qRT-PCR) analysis indicated that the 17 CDPKs were responsive after different degrees of induction under drought and salt stresses. GmCDPK3 was selected as a further research target due to its high relative expression. The subcellular localization experiment showed that GmCDPK3 was located on the membrane of Arabidopsis mesophyll protoplasts. Overexpression of GmCDPK3 improved drought and salt resistance in Arabidopsis. In the soybean hairy roots experiment, the leaves of GmCDPK3 hairy roots with RNA interference (GmCDPK3-RNAi) soybean lines were more wilted than those of GmCDPK3 overexpression (GmCDPK3-OE) soybean lines after drought and salt stresses. The trypan blue staining experiment further confirmed that cell membrane damage of GmCDPK3-RNAi soybean leaves was more severe than in GmCDPK3-OE soybean lines. In addition, proline (Pro) and chlorophyll contents were increased and malondialdehyde (MDA) content was decreased in GmCDPK3-OE soybean lines. On the contrary, GmCDPK3-RNAi soybean lines had decreased Pro and chlorophyll content and increased MDA. The results indicate that GmCDPK3 is essential in resisting drought and salt stresses.

A rice really interesting new gene H2-type E3 ligase, OsSIRH2-14, enhances salinity tolerance via ubiquitin/26S proteasome-mediated degradation of salt-related proteins.

Salinity is a deleterious abiotic stress factor that affects growth, productivity, and physiology of crop plants. Strategies for improving salinity tolerance in plants are critical for crop breeding programmes. Here, we characterized the rice (Oryza sativa) really interesting new gene (RING) H2-type E3 ligase, OsSIRH2-14 (previously named OsRFPH2-14), which plays a positive role in salinity tolerance by regulating salt-related proteins including an HKT-type Na+ transporter (OsHKT2;1). OsSIRH2-14 expression was induced in root and shoot tissues treated with NaCl. The OsSIRH2-14-EYFP fusion protein was predominately expressed in the cytoplasm, Golgi, and plasma membrane of rice protoplasts. In vitro pull-down assays and bimolecular fluorescence complementation assays revealed that OsSIRH2-14 interacts with salt-related proteins, including OsHKT2;1. OsSIRH2-14 E3 ligase regulates OsHKT2;1 via the 26S proteasome system under high NaCl concentrations but not under normal conditions. Compared with wild type plants, OsSIRH2-14-overexpressing rice plants showed significantly enhanced salinity tolerance and reduced Na+ accumulation in the aerial shoot and root tissues. These results suggest that the OsSIRH2-14 RING E3 ligase positively regulates the salinity stress response by modulating the stability of salt-related proteins.

Semi-dominant mutation in the cysteine-rich receptor-like kinase gene, ALS1, conducts constitutive defence response in rice.

Plants have evolved a sophisticated two-branch defence system to prevent the growth and spread of pathogen infection. The novel Cys-rich repeat (CRR) containing receptor-like kinases, known as CRKs, were reported to mediate defence resistance in plants. For rice, there are only two reports of CRKs. A semi-dominant lesion mimic mutant als1 (apoptosis leaf and sheath 1) in rice was identified to demonstrate spontaneous lesions on the leaf blade and sheath. A map-based cloning strategy was used for fine mapping and cloning of ALS1, which was confirmed to be a typical CRK in rice. Functional studies of ALS1 were conducted, including phylogenetic analysis, expression analysis, subcellular location and blast resistance identification. Most pathogenesis-related (PR) genes and other defence-related genes were activated and up-regulated to a high degree. ALS1 was expressed mainly in the leaf blade and sheath, in which further study revealed that ALS1 was present in the vascular bundles. ALS1 was located in the cell membrane of rice protoplasts, and its mutation did not change its subcellular location. Jasmonic acid (JA) and salicylic acid (SA) accumulation were observed in als1, and enhanced blast resistance was also observed. The mutation of ALS1 caused a constitutively activated defence response in als1. The results of our study imply that ALS1 participates in a defence response resembling the common SA-, JA- and NH1-mediated defence responses in rice.

Rational Design Principles for the Transport and Subcellular Distribution of Nanomaterials into Plant Protoplasts.

The ability to control the subcellular localization of nanoparticles within living plants offers unique advantages for targeted biomolecule delivery and enables important applications in plant bioengineering. However, the mechanism of nanoparticle transport past plant biological membranes is poorly understood. Here, a mechanistic study of nanoparticle cellular uptake into plant protoplasts is presented. An experimentally validated mathematical model of lipid exchange envelope penetration mechanism for protoplasts, which predicts that the subcellular distribution of nanoparticles in plant cells is dictated by the particle size and the magnitude of the zeta potential, is advanced. The mechanism is completely generic, describing nanoparticles ranging from quantum dots, gold and silica nanoparticles, nanoceria, and single-walled carbon nanotubes (SWNTs). In addition, the use of imaging flow cytometry to investigate the influence of protoplasts' morphological characteristics on nanoparticle uptake efficiency is demonstrated. Using DNA-wrapped SWNTs as model nanoparticles, it is found that glycerolipids, the predominant lipids in chloroplast membranes, exhibit stronger lipid-nanoparticle interaction than phospholipids, the major constituent in protoplast membrane. This work can guide the rational design of nanoparticles for targeted delivery into specific compartments within plant cells without the use of chemical or mechanical aid, potentially enabling various plant engineering applications.

Morphological characterisation of lettuce plasma membrane ultrastructure and vesicle formation caused by nonylphenol: A scanning electron microscopy study

Fresh produce can come into contact with nonylphenol (NP), a widely used surfactant and industrial endocrine disrupting chemical, through various routes of exposure in the agricultural environment. Nonylphenol has been linked to immunological and behavioural abnormalities as well as infertility and cancer in mammals. One of its modes of action is believed to be through damaging the plasma membrane of cells. While it has been demonstrated that NP can be phytotoxic, its effect on plant cell membrane integrity has not yet been examined. In this study, the plasma membranes of lettuce protoplasts were analysed with a high- resolution scanning electron microscope (SEM). The protoplasts were isolated from plants that had been exposed to 800, 1600, 3200, 6400 and 12,800μg/l NP, respectively, for three weeks. The plasma membrane of the protoplasts that were exposed to 6400 and 12,800μg/l NP showed vesicle-like protrusions, accompanied by flattening of the membrane in its direct vicinity. This study showed that NP caused changes in the ultrastructure of lettuce plasma membranes, thereby compromising its functionality. To the best of our knowledge, this paper describes for the first time the morphology of the plasma membrane using high-resolution SEM.

Exosomes versus microexosomes: Shared components but distinct functions.

In multicellular organisms, cellular components are constantly translocated within cells and are also transported exclusively between limited cells, regardless of their physical distance. Exosomes function as one of the key mediators of intercellular transportation. External vesicles were identified 50years ago in plants and now reconsidered to be exosome-like vesicles. Meanwhile, a well-known exosomal component, tetraspanin CD9, regulates sperm-egg fusion in mammals. A number of Arabidopsis tetraspanins are also expressed in reproductive tissues at fertilization, and are localized at the plasma membrane of protoplasts. Moreover, CD9-containing structures (or 'microexosomes') are released from mouse eggs during their maturation and promote the sperm-egg fusion. This phenomenon implies that two types of shared-component intercellular carriers might be released from multiple types of plant and animal cells, which widely regulate biological phenomena. We herein highlight their discrete structures, formation processes, and functions.

An UDP-Glucosyltransferase Gene from Barley Confers Disease Resistance to Fusarium Head Blight

UDP-glucosyltransferases (UGTs) contribute to Fusarium head blight (FHB) resistance of wheat and barley by glycosylating the deoxynivalenol (DON), which is produced by Fusarium fungus. In this study, seven alleles of barley HvUGT14077 (GenBank No.GU170356.1) were cloned using RT-PCR. Among them, HvUGT-10W1, which was isolated from a FHB resistant barley variety 10W1, was significantly up-regulated in young spikes after F. graminearum (F.g) inoculation. HvUGT-10W1::GFP was subcellularly located in the plasma membrane and cytoplasm of the wheat protoplasts. In vitro antifungal activity assay showed that the HvUGT-10W1 protein exerted obvious inhibition against the growth of F.g. The silencing of the HvUGT-10W1 by virus-induced gene silencing (VIGS) resulted in compromised FHB resistance of 10W1, which was shown by the increased infected colonies on the leaves. These indicated that the barley HvUGT-10W1 may also contribute to F.g resistance in barley and provided a potential candidate gene to develop transgenic barley with enhanced FHB resistance.

Light microscopic observation of acid degradation on cell coat system of Arabidopsis thaliana leaves

This article mainly focuses on the degradation course of enzymatic reaction of cell coat system including cell wall and cytoplasma membrane of the Arabidopsis thaliana leaves under acid conditions. Under light microscopic observation, the cell walls of the leaves of A. thaliana which is under the acid condition are intact and arranged in order at first, thereafter the cell wall begins to degrade partly, and the cells tend to disordered state gradually, afterwards the cell walls are totally degraded. Protoplasts isolated by enzyme digestion totally enter the free state, and accompanied by the degradation of the cytoplasma membrane of the free protoplasts, having overflowed, the organelle surround the nucleus, and thus a new protoplast is reconstituted. We further observed the change of pH in the cell in this course. The result indicated that the cell tended to the pH value decrease during the course of the acid enzymatic degradation, and the pH of the protoplast reconstituted from the organelle came back to its normal level. In conclusion, the acid environment had an important effect on the degradation of the cell coat system of A. thaliana leaves.