A class III peroxidase specifically expressed in pathogen-attacked barley epidermis contributes to basal resistance

Higher plants possess large multigene families encoding secreted class III peroxidase (Prx) proteins. In barley, two Prx cDNAs encoding HvPrx07 and HvPrx08 have been isolated and characterized to some extent with respect to a resistance-mediating function upon attack by the powdery-mildew fungus Blumeria graminis f.sp. hordei ( Bgh ). Here we present evidence for the tissue-specific accumulation of a new Prx mRNA, HvPrx40 , in Bgh -attacked epidermis of barley ( Hordeum vulgare ). The encoded protein is predicted to be secreted into the apoplastic space of epidermal cells due to the absence of a C-terminal extension, which distinguishes it from other Prx proteins reported to accumulate in leaf epidermis. Transient overexpression of HvPrx40 enhanced the resistance of wheat ( Triticum aestivum ) and barley against Blumeria graminis f.sp. tritici (wheat powdery mildew) and Bgh , respectively. These findings were complemented by transient-induced gene silencing showing hypersusceptibility of barley leaf epidermal cells to Bgh . The local accumulation of oxidized 3,3-diaminobenzidine that reflects H₂O₂ production at sites of attempted fungal penetration was not reduced in HvPrx40 -silenced cells, suggesting a role of this peroxidase other than the production of reactive oxygen species.     

三个小麦春化基因的时空表达特性分析(英文)

为明确小麦春化基因的时空表达特性,以中国春和洛旱2号小麦品种为试验材料,利用半定量RT-PCR技术,分析了3个春化基因VERNALIZATION1(VRN1)、VRN2和VRN3的时空表达特性。结果表明,VRN1在中国春的三叶期叶片和根、灌浆期的茎秆和旗叶、花药、胚珠和发育的种子中均有不同程度的表达。在开花前,表达水平呈上升趋势,而花后呈降低的趋势,在干种子和萌发种子的胚芽中没有检测到表达;在洛旱2号中,除了在三叶期的叶片和根中没有检测到表达外,VRN1的表达特性与中国春有相同的趋势。VRN2只在三叶期的叶片和萌发种子的胚芽中表达,在其他检测的组织中没有表达;VRN3的表达与VRN1的时空表达特性相似,但在根中未检测到表达。这一结果为进一步分析普通小麦品种春化发育的分子调控机理提供了重要信息。     

小麦抗白粉病相关基因的转化

利用玉米花青素苷合成调节基因C1-Lc作为报告基因, 通过瞬间表达后愈伤组织表面红色斑点的统计分析, 优化了小麦幼胚愈伤组织的基因枪转化参数。小麦Beclin1类似基因TaTBL和硫代硫酸硫转移酶基因TaTST是2个在白粉菌诱导条件下具有增强表达特性的抗病相关基因。本实验进一步利用基因枪将ubi强启动子控制下的2个基因导入到小麦品种扬麦158的幼胚愈伤组织细胞中, 使用除草剂经两轮选择培养基上的筛选和再生获得抗性植株, 进一步通过抗性植株的PCR分析获得转TaTBL基因植株5株, 转TaTST基因植株6株。转基因植株离体叶片的人工接种实验表明, 外源基因的导入不同程度上增强了植株的白粉病抗性, 表现为延缓了白粉菌的发育。利用玉米花青素苷合成调节基因C1-Lc作为报告基因,通过瞬间表达后愈伤组织表面红色斑点的统计分析,优化了小麦幼胚愈伤组织的基因枪转化参数。小麦Beclin1类似基因TaTBL和硫代硫酸硫转移酶基因TaTST是两个在白粉菌诱导条件下具有增强表达特性的抗病相关基因。本实验进一步利用基因枪将ubi强启动子控制下的两个基因导入到小麦品种扬麦158的幼胚愈伤组织细胞中,使用除草剂经两轮选择培养基上的筛选和再生获得抗性植株,进一步通过抗性植株的PCR分析获得转TaTBL基因植株5株,转TaTST基因植株6株。转基因植株离体叶片的人工接种实验表明,外源基因的导入不同程度上增强了植株的白粉病抗性,表现为延缓了白粉菌的发育。     

A haplotype map of allohexaploid wheat reveals distinct patterns of selection on homoeologous genomes

BackgroundBread wheat is an allopolyploid species with a large, highly repetitive genome. To investigate the impact of selection on variants distributed among homoeologous wheat genomes and to build a foundation for understanding genotype-phenotype relationships, we performed population-scale re-sequencing of a diverse panel of wheat lines.ResultsA sample of 62 diverse lines was re-sequenced using the whole exome capture and genotyping-by-sequencing approaches. We describe the allele frequency, functional significance, and chromosomal distribution of 1.57 million single nucleotide polymorphisms and 161,719 small indels. Our results suggest that duplicated homoeologous genes are under purifying selection. We find contrasting patterns of variation and inter-variant associations among wheat genomes; this, in addition to demographic factors, could be explained by differences in the effect of directional selection on duplicated homoeologs. Only a small fraction of the homoeologous regions harboring selected variants overlapped among the wheat genomes in any given wheat line. These selected regions are enriched for loci associated with agronomic traits detected in genome-wide association studies.ConclusionsEvidence suggests that directional selection in allopolyploids rarely acted on multiple parallel advantageous mutations across homoeologous regions, likely indicating that a fitness benefit could be obtained by a mutation at any one of the homoeologs. Additional advantageous variants in other homoelogs probably either contributed little benefit, or were unavailable in populations subjected to directional selection. We hypothesize that allopolyploidy may have increased the likelihood of beneficial allele recovery by broadening the set of possible selection targets.

Electronic supplementary material

The online version of this article (doi:10.1186/s13059-015-0606-4) contains supplementary material, which is available to authorized users.     

Profiling of short RNAs during fleshy fruit development reveals stage-specific sRNAome expression patterns

Plants feature a particularly diverse population of short (s)RNAs, the central component of all RNA silencing pathways. Next generation sequencing techniques enable deeper insights into this complex and highly conserved mechanism and allow identification and quantification of sRNAs. We employed deep sequencing to monitor the sRNAome of developing tomato fruits covering the period between closed flowers and ripened fruits by profiling sRNAs at 10 time-points. It is known that microRNAs (miRNAs) play an important role in development but very little information is available about the majority of sRNAs that are not miRNAs. Here we show distinctive patterns of sRNA expression that often coincide with stages of the developmental process such as flowering, early and late fruit maturation. Moreover, thousands of non-miRNA sRNAs are differentially expressed during fruit development and ripening. Some of these differentially expressed sRNAs derived from transposons but many derive from protein coding genes or regions that show homology to protein coding genes, several of which are known to play a role in flower and fruit development. These findings raise the possibility of a regulative role of these sRNAs during fruit onset and maturation in a crop species. We also identified six new miRNAs and experimentally validated two target mRNAs. These two mRNAs are targeted by the same miRNA but do not belong to the same gene family, which is rare for plant miRNAs. Expression pattern and putative function of these targets indicate a possible role in glutamate accumulation, which contributes to establishing the taste of the fruit.     

Extracting gene expression patterns and identifying co-expressed genes from microarray data reveals biologically responsive processes

Background  

A common observation in the analysis of gene expression data is that many genes display similarity in their expression patterns and therefore appear to be co-regulated. However, the variation associated with microarray data and the complexity of the experimental designs make the acquisition of co-expressed genes a challenge. We developed a novel method for Extracting microarray gene expression Patterns and Identifying co-expressed Genes, designated as EPIG. The approach utilizes the underlying structure of gene expression data to extract patterns and identify co-expressed genes that are responsive to experimental conditions.     

The human brain has distinct regional expression patterns of estrogen receptor alpha mRNA isoforms derived from alternative promoters

The human estrogen receptor (ER) alpha gene is transcribed from multiple promoters, generating mRNA isoforms with unique 5' ends in the untranslated region. In the present study, alternative promoters were shown to regulate the ERalpha gene expression in different neuronal populations of the human brain. By using in situ hybridization histochemistry, the A and B promoters, but not the C promoter, in the ERalpha gene were found to be active in the human forebrain. The mRNA isoform transcribed from the A promoter was expressed in low levels in most of the brain areas where ERalpha mRNA was present. In contrast, the B promoter mRNA isoform was more restricted, localized predominantly in high-expressing ERalpha mRNA regions. The gross anatomical distribution of the different mRNA isoforms analyzed with RT-PCR generally supported the results obtained by the in situ hybridization. Estrogen is known to modulate many different brain functions, such as neuroendocrine events associated with reproduction, mood, and cognition, likely to be mediated by different neuronal populations. Thus, the current findings of alternative ERalpha promoter expression in distinct neuronal populations suggest that multiple promoter usage is a possible mechanism to achieve differentiated regulation of the ERalpha expression, dependent on the cell phenotype and consequently the functions mediated by the specific neuron.     

Analysis and expression of the class III peroxidase large gene family in Arabidopsis thaliana

Higher plants possess a large set of the classical guaiacol peroxidases (class III peroxidases, E.C. 1.11.1.7). These enzymes have been implicated in a wide array of physiological processes such as H(2)O(2) detoxification, auxin catabolism and lignin biosynthesis and stress response (wounding, pathogen attack, etc.). During the last 10 years, molecular cloning has allowed the isolation and characterization of several genes encoding peroxidases in plants. The achievement of the large scale Arabidopsis genome sequencing, combined with the DNA complementary to RNA (cDNA) expressed sequence tags projects, provided the opportunity to draw up the first comprehensive list of peroxidases in a plant. By screening the available databases, we have identified 73 peroxidase genes throughout the Arabidopsis genome. The evolution of the peroxidase multigene family has been investigated by analyzing the gene structure (intron/exon) in correlation with the phylogenetic relationships between the isoperoxidases. An evolutionary pattern of extensive gene duplications can be inferred and is discussed. Using a cDNA array procedure, the expression pattern of 23 peroxidases was established in the different organs of the plant. All the tested peroxidases were expressed at various levels in roots, while several were also detected in stems, leaves and flowers. The specific functions of these genes remain to be determined.     

Rapid generation of new powdery mildew resistance genes after wheat domestication

Plant defence against pathogens is controlled by disease resistance (R) gene products that directly or indirectly detect specific pathogen effectors. Plant-pathogen interactions have been proposed to follow a co-evolutionary arms-race model where R genes are recent and evolve rapidly in response to structural changes in matching pathogen effectors. However, the longevity and extensive polymorphism of R genes studied were more consistent with balancing selection maintaining ancient and diverse R genes or alleles. In bread wheat (Triticum aestivum), the Pm3 locus confers race-specific resistance to wheat powdery mildew (Blumeria graminis f.sp. triticii). Here we describe recently generated Pm3 resistance alleles that all derive from one susceptible allele, Pm3CS, which is widespread among hexaploid bread-wheat lines. One group of four Pm3 resistance alleles shows few, clearly delimited, polymorphic sequence blocks of ancient origin, embedded in sequences identical to Pm3CS and possibly derived from gene conversion. A second group of three alleles differs from Pm3CS by only two to five mutations, all non-synonymous, and all in the leucine-rich repeat-encoding region. Transient transformation experiments confirmed that Pm3 resistance specificities are based on one or few amino acid changes. The Pm3CS allele was found in wild tetraploid wheat, the ancestor of hexaploid bread wheat, specifically from southern Turkey, a region proposed to be the site of wheat domestication. Based on these data, we propose that the Pm3 resistance alleles were generated in agricultural ecosystems after domestication of wheat 10,000 years ago. The evolution of Pm3 alleles in wheat is best described by the model of evolved recycling, where novel genetic variation is integrated in plant populations together with recycling of old variation.     

Pm50: a new powdery mildew resistance gene in common wheat derived from cultivated emmer

Fungal diseases of wheat, including powdery mildew, cause significant crop, yield and quality losses throughout the world. Knowledge of the genetic basis of powdery mildew resistance will greatly support future efforts to develop and cultivate resistant cultivars. Studies were conducted on cultivated emmer-derived wheat line K2 to identify genes involved in powdery mildew resistance at the seedling and adult plant growth stages using a BC₁ doubled haploid population derived from a cross between K2 and susceptible cultivar Audace. A single gene was located distal to microsatellite marker Xgwm294 on the long arm of chromosome 2A. Quantitative trait loci (QTL) analysis indicated that the gene was also effective at the adult plant stage, explaining up to 79.0 % of the variation in the progeny. Comparison of genetic maps indicated that the resistance gene in K2 was different from Pm4, the only other formally named resistance gene located on chromosome 2AL, and PmHNK54, a gene derived from Chinese germplasm. The new gene was designated Pm50.