Allantoin Alleviates Seed Germination Thermoinhibition in <i>Arabidopsis</i>

Allantoin as the metabolite of purine catabolism can store and remobilize nitrogen for plant growth and development. However, emerging evidence suggests it also contributes to plant tolerance to stress response through altering abscisic acid (ABA) and reducing reactive oxygen species (ROS) level. 1-CYS PEROXIREDOXIN (PER1) is a seed-specific antioxidant that enhances seed longevity through scavenging ROS over-accumulation. High temperature (HT) suppresses seed germination and induces seed secondary dormancy, called as seed germination thermoinhibition. However, the mechanism that allantoin and PER1 regulate seed germination thermoinhibition remains unknown. In this study, we reported that allantoin treatment enhances seed germination under HT stress. Consistently, the aln mutants displayed higher seed germination, as well as more accumulation of endogenous allantoin, than that of wild-type control. Further biochemical and genetic analyses showed that allantoin reduces ABA content under HT, and allantoin targets PER1 to efficiently scavenge HT-induced ROS accumulation, meanwhile, the function of allantoin requires PER1 during seed gemination thermotolerance. Collectively, our finding proposes a novel function of allantoin in enhancing seed germination tolerance to HT, and uncovers the underlying mechanism by which allantoin regulates seed germination through altering ABA metabolism and PER1-mediated ROS level under HT stress.     

Effects of Nano-Cerium Oxide on Seed Germination and Seedling Growth of <i>Vitex negundo</i>

Cerium oxide nanoparticles (CeO₂NPs) are likely to have dramatic impacts on plant performances, yet the effects of CeO₂NPs on seed germination and seedling growth have not been fully explored. In this study, the seed germination and seedling growth of subshrub species Vitex negundo under different concentrations of CeO₂NPs (low-1 mg/L, medium-100 mg/L, high-500 mg/L) have been discussed. Results showed that: (1) The seed germination rate reduces by 11.25% and 2.5% under the low and medium concentrations of CeO₂NPs, respectively, but increased by 7.08% under the high concentration; (2) CeO₂NPs had significant effects on the growth traits (root length, shoot height and biomass) of seedlings, being the highest under the medium concentration and the lowest under the highest concentration; (3) The superoxide dismutase activity was the maximum (355.91 U/g), but the protein concentration was the minimum (3.85 ug/mL) under the high concentration of CeO₂NPs. Our results indicated that the effects of CeO₂NPs on seed germination and seedling growth are concentrationdependency, i.e., low and medium concentrations inhibited while high concentration promoted seed germination, however, seedling growth showed opposite responses. Therefore, appropriate CeO₂NPs concentrations are beneficial to the seed germination and seedling growth of Vitex negundo and improve the physiological performance of seedlings and enhance their adaptability to environmental adversity.     

The BHLH Transcriptional Factor PIF4 Competes with the R2R3-MYB Transcriptional Factor MYB75 to Fine-Tune Seeds Germination under High Glucose Stress

It is known that the high level of sugar including glucose suppresses seed germination through ABA signal. ABI5 is an essential component to mediate ABA-dependent seed germination inhibition, but underlying mechanism needs more investigation. Previous study demonstrated the PIF4 activated the expression of ABI5 to suppress seed germination in darkness. Here we reported that PIF4 also mediated the seed germination inhibition through ABI5 under high concentration of glucose treatment. Furthermore, we found that PIF4 interacted with PAP1, the central factor to control anthocyanin biosynthesis. Such interaction was confirmed in vitro and in planta. Biochemical and physiological analysis revealed that PAP1 bond the promoter of ABI5 to suppress its expression, thus enhanced seed germination under high concentration of glucose treatment. Specially, PAP1 competed with PIF4 to antagonize the activation of PIF4 on ABI5 expression, thus promoted seed germination under high glucose treatment. Given these, we uncover a novel role for PIF4 and PAP1 in controlling seed germination under high glucose treatment, and reveal their antagonistic mechanism by which coordinates ABI5 expression to control seed germination in response to the glucose signal.     

Substrate,Hormone, Winnowing,and Stratification Influence the Seed Germination of <i>Ilex asprella</i> (Hook. et Arn.) Champ. ex Benth

Ilex asprella (Hook. et Arn.) Champ. ex Benth is one of the most important traditional Chinese medicines in southern China. The seeds of Ilex asprella usually have extremely low germination due to their dormancy characteristics, which severely impacts the efficiency of seedling raising and increases labor costs. In this study, to improve the seed germination of I. asprella, the effects of germination substrate, hormone, winnowing, and stratification treatments on the seed germination of I. asprella were investigated. The results of the germination matrix showed that the highest germination percentage of 45.2% was achieved under the 20°C/10°C day/night temperature and vermiculite germination medium conditions. The results of hormone treatments revealed that 100–400 mg/L of gibberellin (GA) and 50–100 mg/L of salicylic acid (SA) were found to be effective in releasing the dormancy of I. asprella seeds. Moreover, winnowing could effectively eliminate unsaturated seeds and impurities, thus improving the seed germination of I. asprella. Furthermore, warm temperature (15°C) stratification could expand the temperature range of I. asprella’s seed germination, which was beneficial for seed germination of I. asprella and for seed nursery at room temperature in production practice. The present study obtained a method to break dormancy and increase seed germination in I. asprella, thereby forming a groundwork for improving the efficiency of large-scale planting of I. asprella.     

Genome-Wide Identification,Evolution and Expression Analyses of <i>GA2ox</i> Gene Family in <i>Brassica napus</i> L.

Gibberellin 2-oxidases (GA2ox) are important enzymes that maintain the balance of bioactive GAs in plants. GA2ox genes have been identified and characterized in many plants, but these genes were not investigated in Brassica napus. Here, we identified 31 GA2ox genes in B. napus and 15 of these BnaGA2ox genes were distributed in the A and C subgenomes. Subcellular localization predictions suggested that all BnaGA2ox proteins were localized in the cytoplasm, and gene structure analysis showed that the BnaGA2ox genes contained 2–4 exons. Phylogenetic analysis indicated that BnGA2ox family proteins in monocotyledons and dicotyledons can be divided into four groups, including two C₁₉-GA2ox and two C₂₀-GA2ox clades. Group 4 is a C₂₀-GA2ox Class discovered recently. Most BnaGA2ox genes had a syntenic relationship with AtGA2ox genes. BnaGA2ox genes in the C subgenome had experienced stronger selection pressure than genes in the A subgenome. BnaGA2ox genes were highly expressed in specific tissues such as those involved in growth and development, and most of them were mainly involved in abiotic responses, regulation of phytohormones and growth and development. Our study provided a valuable evolutionary analysis of GA2ox genes in monocotyledons and dicotyledons, as well as an insight into the biological functions of GA2ox family genes in B. napus.     

<i>DWARF</i> and <i>SMALL SEED1</i>, a Novel Allele of <i>OsDWARF</i>, Controls Rice Plant Architecture,Seed Size,and Chlorophyll Biosynthesis

Plant architecture is a vital agronomic trait to control yield in rice (Oryza sativa L.). A dwarf and small seed 1 (dss1) mutant were obtained from the ethyl methanesulfonate (EMS) mutagenized progeny of a Guizhou glutinous landrace cultivar, Lipingzabianhe. The dss1 mutant displayed phenotypes similar to those of brassinosteroid (BR) deficient mutants, such as dwarfing, dark green and rugose erect leaves, small seeds, and loner neck internode panicles with primary branching. In our previous study, the underlying DSS1 gene was isolated, a novel allele of OsDWARF (OsBR6ox) that encodes a cytochrome P450 protein involved in the BR biosynthetic pathway by MutMap technology. In this work, we confirmed that a Thr335Ile amino acid substitution residing in DSS1/OsDWARF was responsible for the dwarf, panicle architecture, and small seed phenotypes in the dss1 mutants by genetic transformation experiments. The overexpression of OsDWARF in the dss1 mutant background could not only recover dss1 to the normal plant height and panicle architecture but also rescued normal leaf angles, seed size, and leaf color. Thus, the specific mutation in DSS1/OsDWARF influenced plant architecture, seed size, and chlorophyll biosynthesis.     

Estimation of Growth and Photosynthetic Performance of Two C₄ Species (<i>Pennisetum spicatum</i> (L.) Körn. and <i>Zea mays</i> L.) under a Low Temperature Treatment

Pearl millet (Pennisetum spicatum (L.) Körn.) and maize (Zea mays L.) are C₄ grass species grown for feeding humans and animals in Almadinah Almunawwarah, which is in the western part of Saudi Arabia. During the winter, the mean temperature, which drops to 14°C, represents a major problem for the growth of these species in this region. Therefore, the objectives of this research were to investigate the growth response and the photosynthetic performance of P. spicatum and Z. mays under a low temperature stress. The treatments involved daytime and nighttime temperatures of 14/12°C (low temperature) and 24/22°C (optimum temperature). The results indicated that low temperature significantly reduced all growth and physiological parameters, including seed germination, leaf expansion, leaf area, shoot length and root length of the two species compared to those of the control. Additionally, the low temperature significantly decreased the light-saturated assimilation rate (A_sat), quantum yield (ϕ), saturated rate of carbon dioxide uptake (A_max) and efficiency of carboxylation on both species compared to those of the control. Moreover, the values of Fv/Fm and the chlorophyll contents of both species were significantly reduced by low temperature compared to those of the control. It can be concluded that both species had little tolerance to low temperatures.     

AM Fungi and <i>Piriformospora indica</i> Improve Plant Growth of <i>Pinus elliottii</i> Seedlings

Pinus elliottii is an exotic afforestation pine extensively distributed in southern parts of China. In order to understand whether endophytic fungi can affect seedling growth of P. elliottii, Piriformospora indica (Pi), Funnelifcrmis mosseae (Fm), and Diversispora tortuosa (Dt) were inoculated respectively, and the non-inoculated group was set as control. The growth indexes, the contents of soluble sugar and soluble protein, and plant endogenous hormone levels in the leaves of P. elliottii, were analyzed. The results showed that Fm, Dt and Pi colonized the P. elliottii roots to form mycorrhizal structure and chlamydospores arranged in beads respectively. Three fungal inoculants exhibited the stimulated growth responses, whilst Dt illustrated the most positive effect on plant height, single fresh weight, trunk diameter and root system structure, compared with the control. On the other hand, the soluble sugar and soluble protein contents were increased distinctively in mycorrhizal plants. The endogenous IAA, GA3, ZR contents were increased, while the ABA contents were reduced in mycorrhizal plants versus non-mycorrhizal plants. The fungi-induced endogenous hormone changes triggered plant growth improvement of P. elliottii seedlings. This research unraveled the positive effect of AM fungi and P. indica on growth of pine seedlings, while, more application of endophytic fungi to fields needs to be explored.     

Antibacterial Activity and Mechanisms of Ethanol Extracts from <i>Scutellaria baicalensis</i> Georgi and <i>Magnolia officinalis</i> against <i>Phytophthora nicotianae</i>

Phytophthora nicotianae causes substantial economic losses in most countries where tobacco is produced. At present, the control of P. nicotianae mainly depends on chemical methods, with considerable environmental and health issues. We investigated the effects of ethanol extracts from Scutellaria baicalensis Georgi (SBG) and Magnolia officinalis (MO). On mycelial growth, sporangium formation, and zoospore release of P. nicotianae. Both extracts inhibited the growth of P. nicotianae, with mycelial growth inhibition rates of 88.92% and 93.92%, respectively, at 40 mg/mL, and EC50 values of 5.39 and 5.74 mg/mL, respectively. The underlying mechanisms were the inhibition of sporangium formation, the reduction of zoospore number, and the destruction of the mycelium structure. At an SBG extract concentration of 16.17 mg/mL, the inhibition rates for sporangia and zoospores were 98.66% and 99.39%, respectively. At an MO extract concentration of 2.87 mg/mL, the production of sporangia and zoospores was completely inhibited. The hyphae treated with the two plant extracts showed different degrees of deformation and damage. Hyphae treated with SBG extract showed adhesion and local swelling, whereas treatment with MO extract resulted in broken hyphae. Mixture of the extracts resulted in a good synergistic effect.     

Identification and Expression Analysis of Abscisic Acid Signal Transduction Genes in Hemp Seeds

Abscisic acid (ABA) is involved in regulating diverse biological processes, but its signal transduction genes and roles in hemp seed germination are not well known. Here, the ABA signaling pathway members, PYL, PP2C and SnRK2 gene families, were identified from the hemp reference genome, including 7 CsPYL (pyrab-actin resistance1-like, ABA receptor), 8 CsPP2CA (group A protein phosphatase 2c), and 7 CsSnRK2 (sucrose nonfermenting1-related protein kinase 2). The content of ABA in hemp seeds in germination stage is lower than that in non-germination stage. Exogenous ABA (1 or 10 μM) treatment had a significant regulatory effect on the selected PYL, PP2C, SnRK2 gene families. CsAHG3 and CsHAI1 were most significantly affected by exogenous ABA treatment. Yeast two-hybrid experiments were performed to reveal that CsPYL5, CsSnRK2.2, and CsSnRK2.3 could interact with CsPP2CA7 and demonstrate that this interaction was ABA-independent. Our results indicated that CsPYL5, CsSnRK2.2, CsSnRK2.3 and CsPP2CA7 might involve in the ABA signaling transduction pathway of hemp seeds during the hemp seed germination stages. This study suggested that novel genetic views can be brought into investigation of ABA signaling pathway in hemp seeds and lay the foundation for further exploration of the mechanism of hemp seed germination.     

<i>In vitro</i> Germination and Micropropagation of <i>Aconitum vilmorinianum</i>: An Important Medicinal Plant in China

Aconitum vilmorinianum, a well-known traditional Chinese herb, is recently being threatened by overexploitation and environment disturbance. This study was conducted to provide propagation methods through in vitro germination and explant cultivation. Germination was stimulated up to 66.00% on Murashige and Skoog (MS) medium containing 2.0 mg L⁻¹ 6-benzylaminopurine (BAP), 0.1 mg L⁻¹ 1-napthaleneacetic acid (NAA), and 30 g L⁻¹ sucrose. Three bacteria (Pantoea agglomerans, Erwinia persicina, and Pseudomonas tolaasii) would be responsible for consistent contamination during germination. The latter two were effectively eradicated after disinfected. The influence of explant types and hormone combinations on direct and indirect organogenesis was evaluated in the present work. The frequency of shoot induction from axillary bud explants was 100% on the MS fortified with 2.0 mg L⁻¹ BAP and 0.3 mg L⁻¹ NAA. Shoots multiplication was optimized on MS medium supplemented with 0.1 mg L⁻¹ thidiazuron (TDZ) and 0.1 mg L⁻¹ NAA. High callus induction percentage (96.67%) was obtained from stem segments on MS medium with 2.0 mg L⁻¹ 2,4-D, then successfully regenerated into shoots on MS medium in the presence of 0.1 mg L⁻¹ TDZ and 0.2 mg L⁻¹ NAA. The present work could be useful for the utilization and conservation of this valuable species.     

Overexpression of <i>IbSINA5</i> Increases Cold Tolerance through a CBF SINA-COR Mediated Module in Sweet Potato

Seven in absentia (SINA) family proteins play a central role in plant growth, development and resistance to abiotic stress. However, their biological function in plant response to cold stress is still largely unknown. In this work, a seven in absentia gene IbSINA5 was isolated from sweet potato. Quantitative real-time polymerase chain reaction (qRT-PCR) analyses demonstrated that IbSINA5 was ubiquitously expressed in various tissues and organs of sweet potato, with a predominant expression in fibrous roots, and was remarkably induced by cold, drought and salt stresses. Subcellular localization assays revealed that IbSINA5-GFP fusion protein was mainly localized in cytoplasm and nucleus. Overexpression of IbSINA5 in sweet potato led to dramatically improved resistance to cold stress in transgenic plants, which was associated with the up-regulated expression of IbCOR (cold-regulated) genes, increased proline production, and decreased malondialdehyde (MDA) and H₂O₂ accumulation in the leaves of transgenic plants. Furthermore, transient expression of IbCBF3, a C-repeat binding factor (CBF) gene, in the leaf protoplasts of wild type sweet potato plants up-regulated the expression of both IbSINA5 and IbCOR genes. Our results suggest that IbSINA5 could function as a positive regulator in the cold signaling pathway through a CBF-SINA-COR mediated module in sweet potato, and have a great potential to be used as a candidate gene for the future breeding of new plant species with improved cold resistance.     

Titanium Dioxide Nanoparticles Promote Root Growth by Interfering with Auxin Pathways in <i>Arabidopsis thaliana</i>

TiO₂ nanoparticles (nano-TiO₂) are widely used in the world, and a considerable amount of nano-TiO₂ is released into the environment, with toxic effects on organisms. In the various species of higher plants, growth, including seed germination, root elongation, and biomass accumulation, is affected by nano-TiO₂. However, the underlying molecular mechanisms remain to be elucidated. In this study, we observed that nano-TiO₂ promoted root elongation in a dose-dependent manner. Furthermore, we found that nano-TiO₂ elevated auxin accumulation in the root tips of the auxin marker lines DII-VENUS and DR5:: GUS, and, correspondingly, quantitative real-time PCR analysis revealed that nano-TiO₂ increased the expression levels of auxin biosynthesis- and transportrelated genes. GFP fluorescence observation using transgenic PIN2-GFP indicated that nano-TiO₂ promoted root growth by inducing PIN2 accumulation. Thus, we propose that nano-TiO₂ promote root growth in Arabidopsis thaliana by altering the expression levels of auxin biosynthesis- and transport-related genes.     

Differential Responses of <i>NHX1</i> and <i>SOS1</i> Gene Expressions to Salinity in two <i>Miscanthus sinensis</i> Anderss. Accessions with Different Salt Tolerance

The lignocellulosic crop Miscanthus spp. has been identified as a good candidate for biomass production. The responses of Miscanthus sinensis Anderss. to salinity were studied to satisfy the needs for high yields in marginal areas and to avoid competition with food production. The results indicated that the relative advantages of the tolerant accession over the sensitive one under saline conditions were associated with restricted Na⁺ accumulation in shoots. Seedlings of two accessions (salt-tolerant ‘JM0119’ and salt-sensitive ‘JM0099’) were subjected to 0 (control), 100, 200, and 300 mM NaCl stress to better understand the salt-induced biochemical responses of genes involved in Na⁺ accumulation in M. sinensis. The adaptation responses of genes encoding for Na⁺ /H⁺ antiporters, NHX1 and SOS1 to NaCl stress were examined in JM0119 and JM0099.The cDNA sequences of genes examined were highly conserved among the relatives of M. sinensis based on the sequencing on approximate 600 bp-long cDNA fragments obtained from degenerate PCR. These salt-induced variations of gene expression investigated by quantitative real-time PCR provided evidences for insights of the molecular mechanisms of salt tolerance in M. sinensis. The expression of NHX1 was up-regulated by salt stress in JM0119 shoot and root tissues. However, it was hardly affected in JM0099 shoot tissue except for a significant increase at the 100 mM salt treatment, and it was salt-suppressed in the JM0099 root tissue. In the root tissue, the expression of SOS1 was induced by the high salt treatment in JM0119 but repressed by all salt treatments in JM0099. Thus, the remarkably higher expression of NHX1 and SOS1 were associated with the resistance to Na⁺ toxicity by regulation of the Na⁺ influx, efflux, and sequestration under different salt conditions.     

DDG1 and G Protein α Subunit RGA1 Interaction Regulates Plant Height and Senescence in Rice (<i>Oryza sativa</i>)

Many studies have already shown that dwarfism and moderate delayed leaf senescence positively impact rice yield, but the underlying molecular mechanism of dwarfism and leaf senescence remains largely unknown. Here, using map-based cloning, we identified an allele of DEP2, DDG1, which controls plant height and leaf senescence in rice. The ddg1 mutant displayed dwarfism, short panicles, and delayed leaf senescence. Compared with the wild-type, ddg1 was insensitive to exogenous gibberellins (GA) and brassinolide (BR). DDG1 is expressed in various organs, especially in stems and panicles. Yeast two-hybrid assay, bimolecular fluorescent complementation and luciferase complementation image assay showed that DDG1 interacts with the α-subunit of the heterotrimeric G protein. Disruption of RGA1 resulted in dwarfism, short panicles, and darker-green leaves. Furthermore, we found that ddg1 and the RGA1 mutant was more sensitive to salt treatment, suggesting that DDG1 and RGA1 are involved in regulating salt stress response in rice. Our results show that DDG1/DEP2 regulates plant height and leaf senescence through interacting with RGA1.     

Phenotypic Variation among and within Three Peppers Species (<i>Capsicum</i>) from Mexico

The phenotypic variation and its distribution among species, morphotypes, and among and within populations was estimated in 71 populations pertaining to 15 morphotypes of three domesticated species of Capsicum from Mexico. Collections were made in the states of Sinaloa, Nayarit, Oaxaca, Tabasco, and Yucatán in two agroecosystems: Backyard-garden and monoculture. Fifteen phenotypic characteristics were analyzed through one-way variance analysis and multivariate analyses of principal components analysis (PCA) and hierarchical clustering using Ward’s method. The study was performed in a uniform greenhouse experiment. A high variation was found among and within populations in all the measured characteristics. Of the total variation, 13.0% was distributed among species, 27.9% among morphotypes, 8.1% among populations, and 51.0% within populations. Because plants grew in a uniform environment, these results indicate that the differences observed among and within species, morphotypes, and populations have a genetic basis. Univariate and multivariate analyses clearly differentiated morphotypes, suggesting that the category of morphotypes must be used to nominate the infraspecific variation in the domesticated Capsicum. The principal components analysis identified a total of 15 principal components that contributed to explain the total variation. The first two components explained 59.64% of the total variation and seven components explained more than 90% of the total variation. Among the measured characteristics, number of seed per fruit, weight of the fruit, width of the fruit, length of the fruit, stem diameter, days to flowering, and height of the plant contributed to component 1 variation, whereas width of the leaf, length of the leaf, and number of locules, number of fruits and number of seeds per plant contributed to component 2. The hierarchical clustering separated the populations and the morphotypes in two large different groups. One group consisted of populations collected in monoculture conditions and the other group corresponded to population collected from backyard-garden conditions. The monoculture populations were characterized mainly by their longer, wider, and higher weight fruits, plants were of less height, had smaller stem diameters, and lower number of fruits than the populations collected from the backyard-garden conditions. The backyard-garden populations of the Capsicum annuum and Capsicum frutescens species, considered wild or semi-domesticated, constituted a non-differentiated phenotypic group that does not allow dividing them in different species.     

Genome-Wide Identification and Expression Profiling Suggest that Invertase Genes Function in Silique Development and the Response to <i>Sclerotinia sclerotiorum</i> in <i>Brassica napus</i>

Invertase (INV), a key enzyme in sucrose metabolism, irreversibly catalyzes the hydrolysis of sucrose to glucose and fructose, thus playing important roles in plant growth, development, and biotic and abiotic stress responses. In this study, we identified 27 members of the BnaINV family in Brassica napus. We constructed a phylogenetic tree of the family and predicted the gene structures, conserved motifs, cis-acting elements in promoters, physicochemical properties of encoded proteins, and chromosomal distribution of the BnaINVs. We also analyzed the expression of the BnaINVs in different tissues and developmental stages in the B. napus cultivar Zhongshuang 11 using qRT-PCR. In addition, we analyzed RNA-sequencing data to explore the expression patterns of the BnaINVs in four cultivars with different harvest indices and in plants inoculated with the pathogenic fungus Sclerotinia sclerotiorum. We used WGCNA (weighted coexpression network analysis) to uncover BnaINVregulatory networks. Finally, we explored the expression patterns of several BnaINV genes in cultivars with long (Zhongshuang 4) and short (Ningyou 12) siliques. Our results suggest that BnaINVs play important roles in the growth and development of rapeseed siliques and the defense response against pathogens. Our findings could facilitate the breeding of high-yielding B. napus cultivars with strong disease resistance.