Cereal Crop Plants Research Articles

The SULTR gene family in maize (Zea mays L.): Gene cloning and expression analyses under sulfate starvation and abiotic stress

Sulfur is an essential macronutrient required for plant growth, development and stress responses. The family of sulfate transporters (SULTRs) mediates the uptake and translocation of sulfate in higher plants. However, basic knowledge of the SULTR gene family in maize (Zea mays L.) is scarce. In this study, a genome-wide bioinformatic analysis of SULTR genes in maize was conducted, and the developmental expression patterns of the genes and their responses to sulfate starvation and abiotic stress were further investigated. The ZmSULTR family includes eight putative members in the maize genome and is clustered into four groups in the phylogenetic tree. These genes displayed differential expression patterns in various organs of maize. For example, expression of ZmSULTR1;1 and ZmSULTR4;1 was high in roots, and transcript levels of ZmSULTR3;1 and ZmSULTR3;3 were high in shoots. Expression of ZmSULTR1;2, ZmSULTR2;1, ZmSULTR3;3, and ZmSULTR4;1 was high in flowers. Also, these eight genes showed differential responses to sulfate deprivation in roots and shoots of maize seedlings. Transcript levels of ZmSULTR1;1, ZmSULTR1;2, and ZmSULTR3;4 were significantly increased in roots during 12-day-sulfate starvation stress, while ZmSULTR3;3 and ZmSULTR3;5 only showed an early response pattern in shoots. In addition, dynamic transcriptional changes determined via qPCR revealed differential expression profiles of these eight ZmSULTR genes in response to environmental stresses such as salt, drought, and heat stresses. Notably, all the genes, except for ZmSULTR3;3, were induced by drought and heat stresses. However, a few genes were induced by salt stress. Physiological determination showed that two important thiol-containing compounds, cysteine and glutathione, increased significantly under these abiotic stresses. The results suggest that members of the SULTR family might function in adaptations to sulfur deficiency stress and adverse growing environments. This study will lay a foundation for better understanding the functional diversity of the SULTR family and exploring genes of interest for genetic improvement of sulfur use efficiency in cereal crop plants.

Photoperiod-H1 (Ppd-H1) Controls Leaf Size

Leaf size is a major determinant of plant photosynthetic activity and biomass; however, it is poorly understood how leaf size is genetically controlled in cereal crop plants like barley (Hordeum vulgare). We conducted a genome-wide association scan for flowering time, leaf width, and leaf length in a diverse panel of European winter cultivars grown in the field and genotyped with a single-nucleotide polymorphism array. The genome-wide association scan identified PHOTOPERIOD-H1 (Ppd-H1) as a candidate gene underlying the major quantitative trait loci for flowering time and leaf size in the barley population. Microscopic phenotyping of three independent introgression lines confirmed the effect of Ppd-H1 on leaf size. Differences in the duration of leaf growth and consequent variation in leaf cell number were responsible for the leaf size differences between the Ppd-H1 variants. The Ppd-H1-dependent induction of the BARLEY MADS BOX genes BM3 and BM8 in the leaf correlated with reductions in leaf size and leaf number. Our results indicate that leaf size is controlled by the Ppd-H1- and photoperiod-dependent progression of plant development. The coordination of leaf growth with flowering may be part of a reproductive strategy to optimize resource allocation to the developing inflorescences and seeds.

Identification and characterization of the ergochrome gene cluster in the plant pathogenic fungus Claviceps purpurea.

Background Claviceps purpurea is a phytopathogenic fungus infecting a broad range of grasses including economically important cereal crop plants. The infection cycle ends with the formation of the typical purple-black pigmented sclerotia containing the toxic ergot alkaloids. Besides these ergot alkaloids little is known about the secondary metabolism of the fungus. Red anthraquinone derivatives and yellow xanthone dimers (ergochromes) have been isolated from sclerotia and described as ergot pigments, but the corresponding gene cluster has remained unknown. Fungal pigments gain increasing interest for example as environmentally friendly alternatives to existing dyes. Furthermore, several pigments show biological activities and may have some pharmaceutical value.ResultsThis study identified the gene cluster responsible for the synthesis of the ergot pigments. Overexpression of the cluster-specific transcription factor led to activation of the gene cluster and to the production of several known ergot pigments. Knock out of the cluster key enzyme, a nonreducing polyketide synthase, clearly showed that this cluster is responsible for the production of red anthraquinones as well as yellow ergochromes. Furthermore, a tentative biosynthetic pathway for the ergot pigments is proposed. By changing the culture conditions, pigment production was activated in axenic culture so that high concentration of phosphate and low concentration of sucrose induced pigment syntheses.ConclusionsThis is the first functional analysis of a secondary metabolite gene cluster in the ergot fungus besides that for the classical ergot alkaloids. We demonstrated that this gene cluster is responsible for the typical purple-black color of the ergot sclerotia and showed that the red and yellow ergot pigments are products of the same biosynthetic pathway. Activation of the gene cluster in axenic culture opened up new possibilities for biotechnological applications like the dye production or the development of new pharmaceuticals.Electronic supplementary materialThe online version of this article (doi:10.1186/s40694-016-0020-z) contains supplementary material, which is available to authorized users.

A Tandem Array of ent-Kaurene Synthases in Maize with Roles in Gibberellin and More Specialized Metabolism.

While most commonly associated with its role in gibberellin phytohormone biosynthesis, ent-kaurene also serves as an intermediate in more specialized diterpenoid metabolism, as exemplified by the more than 800 known derived natural products. Among these are the maize kauralexins. However, no ent-kaurene synthases (KSs) have been identified from maize. The maize gibberellin-deficient dwarf-5 (d5) mutant has been associated with a loss of KS activity. The relevant genetic lesion has been previously mapped, and was found here to correlate with the location of the KS-like gene ZmKSL3. Intriguingly, this forms part of a tandem array with two other terpene synthases (TPSs). Although one of these, ZmTPS1, has been previously reported to encode a sesquiterpene synthase, and both ZmTPS1 and that encoded by the third gene, ZmKSL5, have lost the N-terminal γ-domain prototypically associated with KS(L)s, all three genes fall within the KS(L) or TPS-e subfamily. Here it is reported that all three genes encode enzymes that are targeted to the plastid in planta, where diterpenoid biosynthesis is initiated, and which all readily catalyze the production of ent-kaurene. Consistent with the closer phylogenetic relationship of ZmKSL3 with previously identified KSs from cereals, only transcription of this gene is affected in d5 plants. On the other hand, the expression of all three of these genes is inducible, suggesting a role in more specialized metabolism, such as that of the kauralexins. Thus, these results clarify not only gibberellin phytohormone, but also diterpenoid phytoalexin biosynthesis in this important cereal crop plant.

Breeding healthy cereals: genetic improvement ofFusarium resistance and consequences for mycotoxins

Although it is generally agreed that increased genetic resistance to fungal colonisation byFusarium head blight (FHB) should lead to reduced damage and particularly to reduced contamination due to mycotoxins, this review aims to highlight on this relation based on published literature in comparison to our own results. We focus here on the major cereal crop plants wheat and barley. Generally, correlations between measures for disease severity on the plants or the seeds and toxin content were found positive and significant. Breeding of new cultivars with reducedFusarium disease severity will therefore lead to a correlated selection response in the direction of reduced toxin contamination, for the prevalent toxins such as deoxynivalenol, but also for less abundant mycotoxins and masked mycotoxins. Choosing resistant cultivars is possibly the best option for mycotoxin reduction right at the beginning of the cereal production chain: on the farmer’s field. Therefore, investment in breeding FHB resistant cultivars will contribute to sustainable reduction of the mycotoxin problems in the cereal production chain, and increase food and feed safety.

Genome-wide cataloging and analysis of alternatively spliced genes in cereal crops.

BackgroundProtein functional diversity at the post-transcriptional level is regulated through spliceosome mediated pre-mRNA alternative splicing (AS) events and that has been widely demonstrated to be a key player in regulating the functional diversity in plants. Identification and analysis of AS genes in cereal crop plants are critical for crop improvement and understanding regulatory mechanisms.ResultsWe carried out the comparative analyses of the functional landscapes of the AS using the consensus assembly of expressed sequence tags and available mRNA sequences in four cereal plants. We identified a total of 8,734 in Oryza sativa subspecies (ssp) japonica, 2,657 in O. sativa ssp indica, 3,971 in Sorghum bicolor, and 10,687 in Zea mays AS genes. Among the identified AS events, intron retention remains to be the dominant type accounting for 23.5 % in S. bicolor, and up to 55.8 % in O. sativa ssp indica. We identified a total of 887 AS genes that were conserved among Z. mays, S. bicolor, and O. sativa ssp japonica; and 248 AS genes were found to be conserved among all four studied species or ssp. Furthermore, we identified 53 AS genes conserved with Brachypodium distachyon. Gene Ontology classification of AS genes revealed functional assignment of these genes in many biological processes with diverse molecular functions.ConclusionsAS is common in cereal plants. The AS genes identified in four cereal crops in this work provide the foundation for further studying the roles of AS in regulation of cereal plant growth and development. The data can be accessed at Plant Alternative Splicing Database (http://proteomics.ysu.edu/altsplice/).Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1914-5) contains supplementary material, which is available to authorized users.

Dormancy release and germination of Echinochloa crus-galli grains in relation to galactomannan-hydrolysing enzyme activity

Echinochloa crus-galli, one of the world's most serious weeds, influences seriously the yield and quality of cereal crop plant. It is well known that E. crus-galli grain is dormant, but its dormant type, as well as its dormancy release and germination in relation to galactomannan-hydrolysing enzyme activity were poorly understood. The cooperation of endo-β-mannanase (EC 3.2.1.78), β-mannosidase (EC 3.2.1.25) and α-galactosidase (EC 3.2.1.22) can hydrolyze the cell walls rich in mannan-based polymers. In the present paper, the mature grains are used as experimental materials, we investigated the water uptake of grains, the effect of removing partial endosperm, after-ripening, stratification and phytohormone on grain germination, and the change in endo-β-mannanase, β-mannosidase and α-galactosidase activities of grains during after-ripening and germination. The results showed that the freshly-collected grains were water-permeable and had only phase I and II of water uptake, while the grains after-ripened for 150 d had an obvious phase III of water uptake. In alternating photoperiod, the germination of grains freshly-collected was zero at 10–35°C, and that of half grains was 11% at 20°C only. The grain germination was notably promoted by after-ripening and stratification, but not by gibberellic acid and 6-benzyladenine at 0.0001–1 mmol L−1. β-Mannosidase activity increased during 0 to 300 d of after-ripening and then decreased. The activity of endo-β-mannanase and α-galactosidase of grains decreased with after-ripening. During grain germination, endo-β-mannanase and β-mannosidase activities obviously increased, while α-galactosidase activity decreased. Our data showed that E. crus-galli grain was a deep physiological dormant, the dormancy release by after-ripening was related to an increasing β-mannosidase activity, and its germination was closely associated with an increasing endo-β-mannanase and β-mannosidase activity; which have provided new knowledge to decrease the harm of E. crus-galli on production of cereal crop plant.

Utilization of a high-throughput shoot imaging system to examine the dynamic phenotypic responses of a C4 cereal crop plant to nitrogen and water deficiency over time.

The use of high-throughput phenotyping systems and non-destructive imaging is widely regarded as a key technology allowing scientists and breeders to develop crops with the ability to perform well under diverse environmental conditions. However, many of these phenotyping studies have been optimized using the model plant Arabidopsis thaliana. In this study, The Plant Accelerator(®) at The University of Adelaide, Australia, was used to investigate the growth and phenotypic response of the important cereal crop, Sorghum bicolor L. Moench and related hybrids to water-limited conditions and different levels of fertilizer. Imaging in different spectral ranges was used to monitor plant composition, chlorophyll, and moisture content. Phenotypic image analysis accurately measured plant biomass. The data set obtained enabled the responses of the different sorghum varieties to the experimental treatments to be differentiated and modelled. Plant architectural instead of architecture elements were determined using imaging and found to correlate with an improved tolerance to stress, for example diurnal leaf curling and leaf area index. Analysis of colour images revealed that leaf 'greenness' correlated with foliar nitrogen and chlorophyll, while near infrared reflectance (NIR) analysis was a good predictor of water content and leaf thickness, and correlated with plant moisture content. It is shown that imaging sorghum using a high-throughput system can accurately identify and differentiate between growth and specific phenotypic traits. R scripts for robust, parsimonious models are provided to allow other users of phenomic imaging systems to extract useful data readily, and thus relieve a bottleneck in phenotypic screening of multiple genotypes of key crop plants.

Red card for pathogens: phytoalexins in sorghum and maize.

Cereal crop plants such as maize and sorghum are constantly being attacked by a great variety of pathogens that cause large economic losses. Plants protect themselves against pathogens by synthesizing antimicrobial compounds, which include phytoalexins. In this review we summarize the current knowledge on phytoalexins produced by sorghum (luteolinidin, apigeninidin) and maize (zealexin, kauralexin, DIMBOA and HDMBOA). For these molecules, we highlight biosynthetic pathways, known intermediates, proposed enzymes, and mechanisms of elicitation. Finally, we discuss the involvement of phytoalexins in plant resistance and their possible application in technology, medicine and agriculture. For those whose world is round we tried to set the scene in the context of a hypothetical football game in which pathogens fight with phytoalexins on the different playing fields provided by maize and sorghum.

Members of the barley NAC transcription factor gene family show differential co-regulation with senescence-associated genes during senescence of flag leaves.

The senescence process of plants is important for the completion of their life cycle, particularly for crop plants, it is essential for efficient nutrient remobilization during seed filling. It is a highly regulated process, and in order to address the regulatory aspect, the role of genes in the NAC transcription factor family during senescence of barley flag leaves was studied. Several members of the NAC transcription factor gene family were up-regulated during senescence in a microarray experiment, together with a large range of senescence-associated genes, reflecting the coordinated activation of degradation processes in senescing barley leaf tissues. This picture was confirmed in a detailed quantitative reverse transcription-PCR (qRT-PCR) experiment, which also showed distinct gene expression patterns for different members of the NAC gene family, suggesting a group of ~15 out of the 47 studied NAC genes to be important for signalling processes and for the execution of degradation processes during leaf senescence in barley. Seven models for DNA-binding motifs for NAC transcription factors were designed based on published motifs, and available promoter sequences of barley genes were screened for the motifs. Genes up-regulated during senescence showed a significant over-representation of the motifs, suggesting regulation by the NAC transcription factors. Furthermore, co-regulation studies showed that genes possessing the motifs in the promoter in general were highly co-expressed with members of the NAC gene family. In conclusion, a list of up to 15 NAC genes from barley that are strong candidates for being regulatory factors of importance for senescence and biotic stress-related traits affecting the productivity of cereal crop plants has been generated. Furthermore, a list of 71 senescence-associated genes that are potential target genes for these NAC transcription factors is presented.

Physiological analysis of nitrogen-efficient rice overexpressing alanine aminotransferase under different N regimes

Cereal crop plants have low nitrogen (N) use efficiency, taking up only 30% to 50% of the applied N fertilizers, with the rest having the potential for loss into the environment as N pollution. One way to address this problem is to improve the nitrogen use efficiency of cereal crops using a transgenic approach. We developed alanine aminotransferase overexpressing rice, and we have previously determined that this modification provided an improved nitrogen-use phenotype to the engineered plants. In this study, the transgenic rice were grown in low, medium, and high nitrogen supply, and morphology, plant N levels, enzymatic activity, metabolite levels, and transcriptome response in the roots and shoots at active and maximum tillering at each N level were measured. The transcriptome response was analysed further using MapMan and PageMan to view multiple comparisons. The transgenic rice plants showed improved nitrogen use efficiency at medium and high N supply, but with few significant changes to the amino acid levels or to the transcriptome. The transgenic plants grown in high N showed up-regulation of transcripts associated with photosynthesis, non-melavonate pathway secondary metabolites, protein degradation, and many unknown function transcripts.

Overexpression of a maize E3 ubiquitin ligase gene enhances drought tolerance through regulating stomatal aperture and antioxidant system in transgenic tobacco

Drought is one of the most important limiting factors in crop production. In our previous study, a putative Arabidopsis thaliana SALT- AND DROUGHT-INDUCED RING FINGER1 (AtSDIR1) homolog encoding a RING-finger protein from Zea mays (ZmRFP1) was cloned and its expression pattern and Ub E3 ligase activity were characterized. However, it is uncertain that ZmRFP1 acts as a positive regulator during drought stress. In this study, we further characterized ZmRFP1 in transgenic tobacco to investigate drought tolerance and possible function mechanisms. Overexpression of ZmRFP1 enhanced drought tolerance in tobacco. The transgenic tobacco lines had more closed stomatal pores, higher proline accumulation, but lower levels of malondialdehyde (MDA) when compared with the wild type (WT) under drought stress. Further investigation showed that ZmRFP1 transgenic plants displayed higher SOD and CAT activities, increased NtSOD and NtCAT transcript levels, and decreased reactive oxygen species (ROS) accumulation under drought stress. Taken together, our results demonstrate that ZmRFP1 confers drought stress tolerance in transgenic tobacco not only by increasing the ability to retain water, but also by reducing ROS accumulation and membrane damage through enhancing the antioxidant system. ZmRFP1 might serve as a candidate gene in genetic improvement for drought tolerance engineering in cereal crop plants.

Multiplex Real‐Time PCR Method for Detection and Quantification of Mycotoxigenic Fungi Belonging to Three Different Genera

Cereal crop plants are colonized by many fungal species such as Aspergillus ochraceus and Penicillium verrucosum, which produce ochratoxins, and Fusarium graminearum, which produces trichothecene mycotoxins. A multiplex real-time PCR method using TaqMan probes was developed to simultaneously detect and quantify these mycotoxigenic Fusarium, Penicillium and Aspergillus species in cereal grains. Primers and probes used in this method were designed targeting the trichothecene synthase (Tri5) gene in trichothecene-producing Fusarium, rRNA gene in Penicillium verrucosum, and polyketide synthase gene (Pks) in Aspergillus ochraceus. The method was highly specific in detecting fungal species containing these genes and was sensitive, detecting up to 3 pg of genomic DNA. These PCR products were detectable over five orders of magnitude (3 pg to 30 ng of genomic DNA). The method was validated by evaluating sixteen barley culture samples for the presence of deoxynivalenol (DON) and ochratoxin A (OTA) producing fungi. Among the barley culture samples tested, 9 were positive for Fusarium spp, 5 tested positive for Penicillium spp, and 2 tested positive for Aspergillus spp. Results were confirmed by traditional microbiological methods. These results indicate that DON- and OTA-producing fungi can be detected and quantified in a single reaction tube using this multiplex real-time PCR method. This method would be helpful in detecting and quantifying the mycotoxin producing fungi such as Fusarium, Aspergillus, and Penicillium in cereal grains and cereal-based foods.

Functional characterization of wheat copalyl diphosphate synthases sheds light on the early evolution of labdane-related diterpenoid metabolism in the cereals

Two of the most agriculturally important cereal crop plants are wheat (Triticum aestivum) and rice (Oryza sativa). Rice has been shown to produce a number of diterpenoid natural products as phytoalexins and/or allelochemicals--specifically, labdane-related diterpenoids, whose biosynthesis proceeds via formation of an eponymous labdadienyl/copalyl diphosphate (CPP) intermediate (e.g., the ent-CPP of gibberellin phytohormone biosynthesis). Similar to rice, wheat encodes a number of CPP synthases (CPS), and the three CPS characterized to date (TaCPS1-3) all have been suggested to produce ent-CPP. However, several of the downstream diterpene synthases will only react with CPP intermediate of normal or syn, but not ent, stereochemistry, as described in the accompanying report. Investigation of additional CPS did not resolve this issue, as the only other functional synthase (TaCPS4) also produced ent-CPP. Chiral product characterization of all the TaCPS then established that TaCPS2 uniquely produces normal, rather than ent-, CPP, thus, providing a suitable substrate source for the downstream diterpene synthases. Notably, TaCPS2 is most homologous to the similarly stereochemically differentiated syn-CPP synthase from rice (OsCPS4), while the non-inducible TaCPS3 and TaCPS4 cluster with the rice OsCPS1 required for gibberellin phytohormone biosynthesis, as well as with a barley (Hordeum vulgare) CPS (HvCPS1) that also is characterized here as similarly producing ent-CPP. These results suggest that diversification of labdane-related diterpenoid metabolism beyond the ancestral gibberellins occurred early in cereal evolution, and included the type of stereochemical variation demonstrated here.

Functional characterization of wheat ent-kaurene(-like) synthases indicates continuing evolution of labdane-related diterpenoid metabolism in the cereals

Wheat (Triticum aestivum) and rice (Oryza sativa) are two of the most agriculturally important cereal crop plants. Rice is known to produce numerous diterpenoid natural products that serve as phytoalexins and/or allelochemicals. Specifically, these are labdane-related diterpenoids, derived from a characteristic labdadienyl/copalyl diphosphate (CPP), whose biosynthetic relationship to gibberellin biosynthesis is evident from the relevant expanded and functionally diverse family of ent-kaurene synthase-like (KSL) genes found in rice the (OsKSLs). Herein reported is the biochemical characterization of a similarly expansive family of KSL from wheat (the TaKSLs). In particular, beyond ent-kaurene synthases (KS), wheat also contains several biochemically diversified KSLs. These react either with the ent-CPP intermediate common to gibberellin biosynthesis or with the normal stereoisomer of CPP that also is found in wheat (as demonstrated by the accompanying paper describing the wheat CPP synthases). Comparison with a barley (Hordeum vulgare) KS indicates conservation of monocot KS, with early and continued expansion and functional diversification of KSLs in at least the small grain cereals. In addition, some of the TaKSLs that utilize normal CPP also will react with syn-CPP, echoing previous findings with the OsKSL family, with such enzymatic promiscuity/elasticity providing insight into the continuing evolution of diterpenoid metabolism in the cereal crop plant family, as well as more generally, which is discussed here.

To Gibberellins and Beyond! Insights into the Evolution of Diterpenoid Metabolism

The requisite production of gibberellins as hormones necessary for higher plant growth and development has provided a genetic reservoir of biosynthetic genes whose duplication and divergence is thought to underlie the widespread production of more specialized diterpenoids. These natural products occur throughout the plant kingdom, although examples of such metabolism are found more broadly in fungi and bacteria as well, with a total of almost 7,000 such natural products already known. Here I present insights into the evolution of such diterpenoid natural products biosynthesis that arise from our systematic approach to biochemical investigations of such metabolism. In particular, using rice as a model system, we have significantly increased understanding of the underlying biosynthetic processes. This includes identification of all the responsible diterpene synthases and number of the cytochrome P450 mono‐oxygenases. Our investigations have further led to evidence for plasticity in these enzymes and, hence, the corresponding metabolic pathways in not only rice, but the related cereal crop plant wheat, providing insights into the facility with which more specialized metabolism can be derived from gibberellin biosynthesis.

The OsGEX2 Gene Promoter Confers Sperm Cell Expression in Transgenic Rice

Expression control elements (i.e., promoters) are crucial components required for the genetic engineering of plants, but relatively few well-characterized organ-specific promoters are available. We have characterized the rice Gamete Expressed 2 (OsGEX2) gene promoter in transgenic rice plants. The OsGEX2 gene (Os09g25650) is homologous to Arabidopsis GEX2, a gene that exhibits expression specifically in the gametic cells of Arabidopsis pollen. The OsGEX2 gene transcript was only detected in rice pollen (or mixed tissues containing pollen) and not found in other organs or tissues. Transgenic rice plants containing an OsGEX2 promoter fused to a GUSPlus reporter gene displayed cell type-specific β-glucuronidase enzyme activity localized within the sperm cells of mature rice pollen. This expression pattern was clearly distinct from the uniform reporter gene activity observed in the pollen vegetative cell of transgenic rice carrying the rice Pollen Specific 1 promoter. The sperm cell reporter gene activity in OsGEX2–GUSPlus transgenic plants correlated well with the native gene transcript levels, suggesting that the cis-regulatory elements necessary for this specificity are present in the 1.9-kb OsGEX2 promoter fragment. The OsGEX2 promoter with its cell type-specific expression will be a useful tool for precisely controlling sperm cell gene expression in rice and potentially other cereal crop plants.

ZmRFP1, the putative ortholog of SDIR1, encodes a RING-H2 E3 ubiquitin ligase and responds to drought stress in an ABA-dependent manner in maize

Drought is one of the most important limiting factors in crop production. To identify genes required for the drought stress response in the cereal crop maize, a gene coding for RING-finger protein (ZmRFP1), which is highly responsive to PEG-induced drought stress, was isolated by mRNA differential display and rapid amplification of cDNA ends. The ZmRFP1 encodes a protein of 280 amino acids and contains a single C3H2C3-type RING motif in its C-terminal region. ZmRFP1 is an ortholog of Arabidopsis SDIR1 (salt- and drought-induced RING finger 1) (66% identity to AtSDIR1).The recombinant ZmRFP1 protein purified from Escherichia coli exhibited an in vitro E3 ubiquitin ligase activity. Real-time PCR analysis indicates that the transcript levels of ZmRFP1 were higher in aerial tissues including stems, leaves, tassels and immature ears, and were markedly up-regulated by drought stress, and exogenous ABA, but not by salt, heat and cold stresses. Transient expression of the green fluorescent protein (GFP)-ZmRFP1 fusion protein in onion cells revealed a plasma membrane localization of the protein. Further analysis of ZmRFP1 transcripts between an ABA-deficient transposon mutant viviparous14 (vp14) and its isogenic wild-type line W22 showed that ZmRFP1 transcript levels were induced significantly in the wild-type line under drought stress, but not in the mutant line VP14. These results indicate that ZmRFP1 responds to drought stress in an ABA-dependent way and is likely to function in the ubiquitin conjunction pathway. The ZmRFP1 might serve as a candidate gene in genetic improvement for drought tolerance engineering in cereal crop plants.

Characterization of barley (Hordeum vulgare L.) NAC transcription factors suggests conserved functions compared to both monocots and dicots.

BackgroundThe NAC transcription factor family is involved in the regulation of traits in both monocots and dicots of high agronomic importance. Understanding the precise functions of the NAC genes can be of utmost importance for the improvement of cereal crop plants through plant breeding. For the cereal crop plant barley (Hordeum vulgare L.) only a few NAC genes have so far been investigated.ResultsThrough searches in publicly available barley sequence databases we have obtained a list of 48 barley NAC genes (HvNACs) with 43 of them representing full-length coding sequences. Phylogenetic comparisons to Brachypodium, rice, and Arabidopsis NAC proteins indicate that the barley NAC family includes members from all of the eight NAC subfamilies, although by comparison to these species a number of HvNACs still remains to be identified. Using qRT-PCR we investigated the expression profiles of 46 HvNACs across eight barley tissues (young flag leaf, senescing flag leaf, young ear, old ear, milk grain, late dough grain, roots, and developing stem) and two hormone treatments (abscisic acid and methyl jasmonate).ConclusionsComparisons of expression profiles of selected barley NAC genes with the published functions of closely related NAC genes from other plant species, including both monocots and dicots, suggest conserved functions in the areas of secondary cell wall biosynthesis, leaf senescence, root development, seed development, and hormone regulated stress responses.

Gene silencing goes viral and uncovers the private life of plants

To understand how genes influence plant responses to the environment, it is important to have appropriate tools for studying gene function. This may be a challenge for non-model plant species or plants that are recalcitrant to genetic transformation. Virus-induced gene silencing (VIGS) has been established as a convenient, rapid, and efficient method for reverse genetics in a wide range of plant species ranging from non-model dicot plants to cereal crop plants. Virus-induced gene silencing is not limited to the green parts of a plant; genes can also be efficiently silenced in roots and reproductive organs. As it is established during later developmental stages, it avoids the problem of embryo- or seedling lethality, which may impede the investigation of some null mutants or transgenic knock-out plants. Virus-induced gene silencing can be designed to target specific members of a gene family, or to co-silence paralogous genes, and it has been adapted for high-throughput analysis. Virus-induced gene silencing has frequently been used to study plant–pathogen interactions and plant development. This review emphasizes the great potential of VIGS for the study of plant–arthropod interactions, ranging from plant–herbivore and plant–pollinator interactions to behavioral animal responses to plants. In spite of this potential, VIGS has not often been adopted by entomologists and plant scientists who study plant–arthropod interactions. We also provide practical considerations for developing or adopting a VIGS system.