Analysis of gene expressions associated with increased allelopathy in rice (Oryza sativa L.) induced by exogenous salicylic acid

The defense characteristics of allelopathic rice accession PI312777 and its counterpart Lemont induced by exogenous salicylic acid (SA) to suppress troublesome weed barnyardgrass (BYG) were investigated using the methods of suppression subtractive hybridization (SSH) and real-time fluorescence quantitative PCR (qRT-PCR). The results showed that exogenous SA could induce the allelopathic effect of rice on BYG and this inducible defense was SA dose-respondent and treatment time-dependent. PI312777 exhibited higher inhibitory effect than Lemont on BYG after treated with different concentrations of SA. The activities of cell protective enzymes including SOD, POD and CAT in the BYG plants co-cultured with PI312777 treated by SA were highly depressed compared with the control (co-cultured with rice without SA-treatment). Similar but lower depression on these enzymes except for CAT was also observed in the BYG plants when co-cultured with Lemont treated by SA. It is therefore suggested that allelopathic rice should be more sensitive than non-allelopathic rice to exogenous SA. Seventeen genes induced by SA were obtained by SSH analysis from PI312777. These genes encode receptor-kinase proteins, ubiquitin carrier proteins, proteins related to phenylpropanoid metabolism, antioxidant related proteins and some growth-mediating proteins. The differential expressions of these genes were validated in part by qRT-PCR in the two rice accessions. Our work elucidated that allelopathic rice possesses an active chemical defense and auto-detoxifying enzyme system such as the up-regulated enzymes involved in de novo biosynthesis of phenolic allelochemicals and the glutathione-S-transferase (GST) associated with xenobiotic detoxification.     

The CarD/CarG regulatory complex is required for the action of several members of the large set of Myxococcus xanthus extracytoplasmic function σ factors

Extracytoplasmic function (ECF) σ factors are critical players in signal transduction networks involved in bacterial response to environmental changes. The Myxococcus xanthus genome reveals ～45 putative ECF‐σ factors, but for the overwhelming majority, the specific signals or mechanisms for selective activation and regulation remain unknown. One well‐studied ECF‐σ, CarQ, binds to its anti‐σ, CarR, and is inactive in the dark but drives its own expression from promoter P_QRS on illumination. This requires the CarD/CarG complex, the integration host factor (IHF) and a specific CarD‐binding site upstream of P_QRS. Here, we show that DdvS, a previously uncharacterized ECF‐σ, activates its own expression in a CarD/CarG‐dependent manner but is inhibited when specifically bound to the N‐terminal zinc‐binding anti‐σ domain of its cognate anti‐σ, DdvA. Interestingly, we find that the autoregulatory action of 11 other ECF‐σ factors studied here depends totally or partially on CarD/CarG but not IHF. In silico analysis revealed possible CarD‐binding sites that may be involved in direct regulation by CarD/CarG of target promoter activity. CarD/CarG‐linked ECF‐σ regulation likely recurs in other myxobacteria with CarD/CarG orthologous pairs and could underlie, at least in part, the global regulatory effect of the complex on M. xanthus gene expression.     

Dual targeting of Myxococcus xanthus protoporphyrinogen oxidase into chloroplasts and mitochondria and high level oxyfluorfen resistance

Much attention has been paid to the signal sequences of eukaryotic protoporphyrinogen oxidases (protoxes); both the organelles targeted by protoxes and the role of protoxes in conferring resistance against protox‐inhibiting herbicides, such as oxyfluorfen, have been examined. However, there have been no reports on the translocation of prokaryotic protoxes. This study investigated the targeting ability of Myxococcus xanthus protox in vitro and in vivo. In an in vitro translocation assay using a dual import system, M. xanthus protein was detected in chloroplasts and mitochondria, suggesting that the M. xanthus protox protein was targeted into both organelles. In order to confirm the in vitro dual targeting ability of M. xanthus, we used a stable transgenic strategy to investigate dual targeting in vivo. In transgenic rice plants overexpressing M. xanthus protox, M. xanthus protox antibody cross‐reacted with proteins with predicted molecular masses of 50 kDa from both chloroplasts and mitochondria, and this in vivo transgene expression corresponded to a prominent increase in chloroplastic and mitochondrial protox activity. Seeds from the transgenic lines M4 and M7 germinated in solid Murashige and Skoog media of up to 500 µm of oxyfluorfen, whereas wild‐type seeds did not germinate in 1 µm . After 4‐week‐old‐rice plants were treated with oxyfluorfen for 3 d, lines M4 and M7 exhibited normal growth, whereas the wild‐type line was severely bleached and necrotized. The herbicidal resistance is attributed to the insignificant accumulation of photodynamic protoporphyrin IX in cytosol because the high chloroplastic and mitochondrial protox activity in oxyfluorfen‐treated transgenic lines, compared with that in oxyfluorfen‐treated and untreated wild‐type plants, metabolizes protoporphyrinogen IX to chlorophyll and heme. A practical application of the dual targeting of M. xanthus protox for obtaining outstanding resistance to peroxidizing herbicides is discussed.     

Identification of microRNAs and their corresponding targets involved in the susceptibility interaction of wheat response to Puccinia striiformis f. sp. tritici

MicroRNAs (miRNAs) play very important roles in plant defense responses. However, little is known about their roles in the susceptibility interaction between wheat and Puccinia striiformis f. sp. tritici (Pst). In this study, two miRNA libraries were constructed from the leaves of the cultivar Xingzi 9104 inoculated with the virulent Pst race CYR32 and sterile water, respectively. A total of 1316 miRNA candidates, including 173 known miRNAs that were generated from 98 pre‐miRNAs, were obtained. The remaining 1143 miRNA candidates included 145 conserved and 998 wheat‐specific miRNAs that were generated from 87 and 1088 pre‐miRNAs, respectively. The 173 known and 145 conserved miRNAs were sub‐classified into 63 miRNA families. The target genes of wheat miRNAs were also confirmed using degradome sequencing technology. Most of the annotated target genes were related to signal transduction or energy metabolism. Additionally, we found that miRNAs and their target genes form complicated regulation networks. The expression profiles of miRNAs and their corresponding target genes were further analyzed by quantitative real‐time polymerase chain reaction (qRT‐PCR), and the results indicate that some miRNAs are involved in the compatible wheat‐Pst susceptibility interaction. Importantly, tae‐miR1432 was highly expressed when wheat was challenged with CYR32, and the corresponding target gene, predicted to be a calcium ion‐binding protein, also exhibited upregulated expression but a divergent expression trend. PC‐3P‐7484, a specific wheat miRNA, was highly expressed in the wheat response to Pst infection, while the expression of the corresponding target gene ubiquillin was dramatically downregulated. These data provide the foundation for evaluating the important regulatory roles of miRNAs in wheat‐Pst susceptibility interaction.     

Interference of allelopathic rice with paddy weeds at the root level

• Despite increasing knowledge of the involvement of allelopathy in negative interactions among plants, relatively little is known about its action at the root level. This study aims to enhance understanding of interactions of roots between a crop and associated weeds via allelopathy.
• Based on a series of experiments with window rhizoboxes and root segregation methods, we examined root placement patterns and root interactions between allelopathic rice and major paddy weeds Cyperus difformis, Echinochloa crus‐galli, Eclipta prostrata, Leptochloa chinensis and Oryza sativa (weedy rice).
• Allelopathic rice inhibited growth of paddy weed roots more than shoots regardless of species. Furthermore, allelopathic rice significantly reduced total root length, total root area, maximum root width and maximum root depth of paddy weeds, while the weeds adjusted horizontal and vertical placement of their roots in response to the presence of allelopathic rice. With the exception of O. sativa (weedy rice), root growth of weeds avoided expanding towards allelopathic rice. Compared with root contact, root segregation significantly increased inhibition of E. crus‐galli, E. prostrata and L. chinensis through an increase in rice allelochemicals. In particular, their root exudates induced production of rice allelochemicals. However, similar results were not observed in C. difformis and O. sativa (weedy rice) with either root segregation or root exudate application.
• The results demonstrate that allelopathic rice interferes with paddy weeds by altering root placement patterns and root interactions. This is the first case of a root behavioural strategy in crop–weed allelopathic interaction.

     

Allelopathy in wheat (Triticum aestivum)

Wheat (Triticum aestivum) allelopathy has potential for the management of weeds, pests and diseases. Both wheat residue allelopathy and wheat seedling allelopathy can be exploited for managing weeds, including resistant biotypes. Wheat varieties differ in allelopathic potential against weeds, indicating that selection of allelopathic varieties might be a useful strategy in integrated weed management. Several categories of allelochemicals for wheat allelopathy have been identified, namely, phenolic acids, hydroxamic acids and short‐chain fatty acids. Wheat allelopathic activity is genetically controlled and a multigenic model has been proposed. Research is underway to identify genetic markers associated with wheat allelopathy. Once allelopathic genes have been located, a breeding programme could be initiated to transfer the genes into modern varieties for weed suppression. The negative impacts of wheat autotoxicity on agricultural production systems have also been identified when wheat straws are retained on the soil surface for conservation farming purposes. A management package to avoid such deleterious effects is discussed. Wheat allelopathy requires further study in order to maximise its allelopathic potential for the control of weeds, pests and diseases, and to minimise its detrimental effects on the growth of wheat and other crops.     

Rice allelopathy and the possibility for weed management

In attempts to control weeds in rice, much effort has been focused on rice allelopathy research for more than 30 years. Among screening methods that have been developed, some estimate the allelopathic potential of various rice cultivars in a limited time and space, which is less costly and can be conducted year round. Rice allelopathy activity is variety dependent and origin dependent, where Japonica rice shows greater allelopathic activity than Indica and Japonica–Indica hybrids. Allelopathic characteristics in rice are quantitatively inherited and several allelopathy‐involved traits have been identified. Numerous phytotoxins such as cytokinins, diterpenoids, fatty acids, flavones, glucopyranosides, indoles, momilactones (A and B), oryzalexins, phenols, phenolic acids, resorcinols and stigmastanols have been identified and determined as growth inhibitors in rice. However, the fate and actual modes of action of these compounds as well as other potent rice phytotoxins in nature are not well understood. The question of which compounds play a major role in rice allelopathy has remained obscure; however, rice allelopathy might be attributable to the interaction of all present allelochemicals. Despite locating genes determining or involving allelopathy in rice having attracted much effort, the introduction of these genes into target rice cultivars has not yet been achieved. Success in breeding new rice cultivars having good weed‐suppressing ability would benefit farmers in rice‐cultivating countries and play an important role in sustainable agricultural production.     

Identification and Analysis of Powdery Mildew‐Responsive miRNAs in Wheat

Wheat powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is one of the most consistently damaging diseases of common wheat worldwide and greatly affects crop productivity. Recently, several plant microRNAs (miRNAs) have been reported as gene expression regulators related to various adverse environments. However, up to now, less is known on the roles of miRNAs in powdery mildew infection response of wheat. In this study, miRNA expression patterns were investigated for identifying Bgt‐responsive miRNAs in wheat leaves using a plant miRNA microarray platform. A total of 79 miRNAs from 24 families were detected in wheat leaves. Among those, seven miRNAs were further validated to be involved in wheat powdery mildew response and two of them have never been reported. In addition, their target expression profiles showed a negative correlation with that of the seven miRNAs in mock‐ and Bgt‐infected samples furtherly proved, which in turn as the robust evidence, that those seven powdery mildew‐responsive miRNAs are highly reliable. These findings could extend the current view about miRNAs as ubiquitous regulators under stress conditions.     

Identification of MicroRNAs controlling human ovarian cell proliferation and apoptosis

Previous studies have shown that microRNAs (miRNAs) can control steroidogenesis in cultured granulosa cells. In this study we wanted to determine if miRNAs can also affect proliferation and apoptosis in human ovarian cells. The effect of transfection of cultured primary ovarian granulosa cells with 80 different constructs encoding human pre‐miRNAs on the expression of the proliferation marker, PCNA, and the apoptosis marker, Bax was evaluated by immunocytochemistry. Eleven out of 80 tested miRNA constructs resulted in stimulation, and 53 miRNAs inhibited expression of PCNA. Furthermore, 11 of the 80 miRNAs tested promoted accumulation of Bax, while 46 miRNAs caused a reduction in Bax in human ovarian cells. In addition, two selected antisense constructs that block the corresponding miRNAs mir‐15a and mir‐188 were evaluated for their effects on expression of PCNA. An antisense construct inhibiting mir‐15a (which precursor suppressed PCNA) increased PCNA, whereas an antisense construct for mir‐188 (which precursor did not change PCNA) did not affect PCNA expression. Verification of effects of selected pre‐mir‐10a, mir‐105, and mir‐182 by using other markers of proliferation (cyclin B1) and apoptosis (TdT and caspase 3) confirmed specificity of miRNAs effects on these processes. This is the first direct demonstration of the involvement of miRNAs in controlling both proliferation and apoptosis by ovarian granulose cells, as well as the identification of miRNAs promoting and suppressing these processes utilizing a genome‐wide miRNA screen. J. Cell. Physiol. 223: 49–56, 2010. © 2009 Wiley‐Liss, Inc.     

Allelochemicals released by rice roots and residues in soil

A few rice (Oryza sativa L.) varieties or rice straw produce and release allelochemicals into soil in which interfere with the growth of neighboring or successive plants. Allelopathic rice PI312777 and Huagan-1 at their early growth stages released momilactone B, 3-isopropyl-5-acetoxycyclohexene-2-one-1, and 5,7,4′-trihydroxy-3′,5′-dimethoxyflavone into soil at phytotoxic levels, but non-allelopathic rice Huajingxian did not. Both allelopathic and non-allelopathic rice residues released momilactone B and lignin-related phenolic acids (p-hydroxybenzoic, p-coumaric, ferulic, syringic and vanillic acids) into the soil during residue decomposition to inhibit successive plants. The results indicated that allelochemicals involved in rice allelopathy from living and dead plants are substantially different. Interestingly, the concentrations of the allelochemicals released from the allelopathic rice seedlings in soil increased dramatically when they were surrounded with Echinochloa crus-galli. The concentrations of the allelochemicals were over 3-fold higher in the presence of E. crus-galli than in the absence of E. crus-galli. However, the same case did not occur in non-allelopathic Huajingxian seedlings surrounded with E. crus-galli. In addition to allelochemical exudation being promoted by the presence of E. crus-galli, allelopathic rice seedlings also increased allelochemical exudation in response to exudates of germinated E. crus-galli seeds or lepidimoide, an uronic acid derivative exuded from E. crus-galli seeds. These results imply that allelopathic rice seedlings can sense certain allelochemicals released by E. crus-galli into the soil, and respond by increased production of allelochemicals inhibitory to E. crus-galli. This study suggests that rice residues of both allelopathic and non-allelopathic varieties release similar concentrations and types of allelochemicals to inhibit successive plants. In contrast, living rice plants of certain allelopathic varieties appear to be able to detect the presence of interspecific neighbors and respond by increased allelochemicals.     

Separation of allelopathy from resource competition using rice/barnyardgrass mixed-cultures

Plant-plant interference is the combined effect of allelopathy, resource competition, and many other factors. Separating allelopathy from resource competition is almost impossible in natural systems but it is important to evaluate the relative contribution of each of the two mechanisms on plant interference. Research on allelopathy in natural and cultivated plant communities has been hindered in the absence of a reliable method that can separate allelopathic effect from resource competition. In this paper, the interactions between allelopathic rice accession PI312777, non-allelopathic rice accession Lemont and barnyardgrass were explored respectively by using a target (rice)-neighbor (barnyardgrass) mixed-culture in hydroponic system. The relative competitive intensity (RCI), the relative neighbor effect (RNE) and the competitive ratio (CR) were used to quantify the intensity of competition between each of the two different potentially allelopathic rice accessions and barnyardgrass. Use of hydroponic culture system enabled us to exclude any uncontrolled factors that might operate in the soil and we were able to separate allelopathy from resource competition between each rice accession and barnyardgrass. The RCI and RNE values showed that the plant-plant interaction was positive (facilitation) for PI312777 but that was negative (competition) for Lemont and barnyardgrass in rice/barnyardgrass mixed-cultures. The CR values showed that one PI312777 plant was more competitive than 2 barnyardgrass plants. The allelopathic effects of PI312777 were much more intense than the resource competition in rice/barnyardgrass mixed cultures. The reverse was true for Lemont. These results demonstrate that the allelopathic effect of PI312777 was predominant in rice/barnyardgrass mixed-cultures. The most significant result of our study is the discovery of an experimental design, target-neighbor mixed-culture in combination with competition indices, can successfully separate allelopathic effects from competition.     

水稻化感作用及其分子生态学研究进展

综述了近年来国际上研究水稻化感作用的新进展,比较分析了当前常用于室内评价水稻化感作用潜力的几种生物测试法的优缺点,指出了琼脂迟播共培法是较为理想的室内生物测试法并已广泛应用于化感作用研究中。在此基础上,分析了水稻化感作用的数量遗传特性及其QTL定位的研究现状;阐明了水稻化感作用的遗传多样性及其分子生态特性;并就当前普遍关注的焦点问题：逆境条件（如低氮或高伴生杂草密度胁迫）常引起水稻化感作用潜力增大的生理过程与分子机制作了阐述。结合近年来应用差异蛋白组学和生物信息学的研究实例,阐明了逆境引起水稻化感作用增强与其酚类合成代谢相关酶蛋白表达丰度增加,萜类合成代谢相关酶蛋白表达丰度下降有关。就究竟什么是水稻的化感物质及其作用方式等问题作了分析与讨论．指出水稻的化感抑草效应是其众多化感物质综合作用的结果,应重视区分化感物质对靶标杂草的原生作用和进入土壤生态系统中经生物转化后的次生作用。根据当前植物化感作用研究的发展趋势,阐明了进一步研究水稻化感作用的焦点问题,提出了水稻化感作用的根际生物学特性与分子生态学机制,是未来国际上竞相角逐的重点研究领域,并认为以现代系统生物学理论为指导,运用基因组学、蛋白质组学和代谢组学等技术方法,是揭示这一分子生态学过程与机制的重要技术选择和优先研究领域。     

Allelopathy prevents competitive exclusion and promotes phytoplankton biodiversity

It has been hypothesized that allelopathy can prevent competitive exclusion and promote phytoplankton diversity in aquatic ecosystems, where numerous species coexist on a limited number of resources. However, experimental proof‐of‐principle is not available to support this hypothesis. Here we present the first experimental evidence to support this hypothesis by demonstrating that allelopathy promotes the coexistence of two phytoplankton species, Ankistrodesmus falcatus and Oscillatoria sp., that compete for a single limiting nutrient. By performing long‐term competition experiments in nitrate‐limited continuous cultures, and by describing the population dynamics using a mechanistic model, we demonstrate that when allelopathy comes into play, one of the following outcomes is possible depending on the relative initial abundances of the species: dominance of the stronger competitor for nitrate (the non‐allelopathic species), oscillatory coexistence, or dominance of the weaker competitor (the allelopathic species). Our model analysis revealed that sustained oscillatory coexistence of the two species would be a common outcome of this experiment. Our study confirms for the first time, based on laboratory experiments combined with mechanistic models, that allelopathy can alter the predicted outcome of inter‐specific competition in a nutrient‐limited environment and increase the potential for the coexistence of more species than resources, thereby contributing to the identification of endogenous mechanisms that explain the extreme diversity of phytoplankton communities.     

Genome‐wide identification and characterization of microRNAs responding to ABA and GA in maize embryos during seed germination

• MicroRNAs (miRNAs) are an important class of non‐coding small RNAs that regulate the expression of target genes through mRNA cleavage or translational inhibition. Previous studies have revealed their roles in regulating seed dormancy and germination in model plants such as Arabidopsis thaliana, rice (Oryza sativa) and maize (Zea mays). However, the miRNA response to exogenous gibberellic acid (GA) and abscisic acid (ABA) during seed germination in maize has yet to be explored.
• In this study, small RNA libraries were generated and sequenced from maize embryos treated with GA, ABA or double‐distilled water as control.
• A total of 247 miRNAs (104 known and 143 novel) were identified, of which 45 known and 53 novel miRNAs were differentially expressed in embryos in the different treatment groups. In total, 74 (37 up‐regulated and 37 down‐regulated) and 55 (23 up‐regulated and 32 down‐regulated) miRNAs were expressed in response to GA and to ABA, respectively, and a total of 18 known and 38 novel miRNAs displayed differential expression between the GA‐ and ABA‐treated groups. Using bioinformatics tools, we predicted the target genes of the differentially expressed miRNAs. Using GO enrichment and KEGG pathway analysis of these targets, we showed that miRNAs differentially expressed in our samples affect genes encoding proteins involved in the peroxisome, ribosome and plant hormonal signalling pathways.
• Our results indicate that miRNA‐mediated gene expression influences the GA and ABA signalling pathways during seed germination.