BackgroundMicroRNAs (miRNAs) are small, non-coding RNAs playing essential roles in plant growth, development, and stress responses. Sequencing of small RNAs is a starting point for understanding their number, diversity, expression and possible roles in plants.ResultsIn this study, we conducted a genome-wide survey of wheat miRNAs from 11 tissues, characterizing a total of 323 novel miRNAs belonging to 276 families in wheat. A miRNA conservation analysis identified 191 wheat-specific miRNAs, 2 monocot-specific miRNAs, and 30 wheat-specific variants from 9 highly conserved miRNA families. To understand possible roles of wheat miRNAs, we determined 524 potential targets for 124 miRNA families through degradome sequencing, and cleavage of a subset of them was validated via 5′ RACE. Based on the genome-wide identification and characterization of miRNAs and their associated target genes, we further identified 64 miRNAs preferentially expressing in developing or germinating grains, which could play important roles in grain development.ConclusionWe discovered 323 wheat novel miRNAs and 524 target genes for 124 miRNA families in a genome-wide level, and our data will serve as a foundation for future research into the functional roles of miRNAs in wheat.

BackgroundIn angiosperms, the endosperm plays a crucial placenta-like role in that not only is it necessary for nurturing the embryo, but also regulating embryogenesis through complicated genetic and epigenetic interactions with other seed compartments and is the primary tissue in which genomic imprinting occurs.ResultsWe observed a gradual increase of paternal siRNA expression in the early stages of kernels and an expected 2:1 maternal to paternal ratio in 7-DAP endosperm via sequencing of small interfering RNA (siRNA) transcriptomes in developing kernels (0, 3 and 5 days after pollination (DAP)) and endosperms (7, 10 and 15 DAP) from the maize B73 and Mo17 reciprocal crosses. Additionally, 460 imprinted siRNA loci were identified in the endosperm, with the majority (456/460, 99.1%) being maternally expressed at 10 DAP. Moreover, 13 out of 29 imprinted genes harbored imprinted siRNA loci within their 2-kb flanking regions, a significant higher frequency than expected based on simulation analysis. Additionally, gene ontology terms of “response to auxin stimulus”, “response to brassinosteroid stimulus” and “regulation of gene expression” were enriched with genes harboring 10-DAP specific siRNAs, whereas those of “nutrient reservoir activity”, “protein localization to vacuole” and “secondary metabolite biosynthetic process” were enriched with genes harboring 15-DAP specific siRNAs.ConclusionsA subset of siRNAs subjected to imprinted expression pattern in maize developing endosperm, and they are likely correlated with certain imprinted gene expression. Additionally, siRNAs might influence nutrient uptake and allocation processes during maize endosperm development.

BackgroundMicroRNAs are a class of small, non-coding RNAs that regulate gene expression by binding target mRNA, which leads to cleavage or translational inhibition. The NAC proteins, which include NAM, ATAF, and CUC, are a plant-specific transcription factor family with diverse roles in development and stress regulation. It has been reported that miR164 negatively regulates NAC1 expression, which in turn affects lateral root development in Arabidopsis; however, little is known about the involvement of the maize NAC family and miR164 in lateral root development.ResultsWe collected 175 maize transcripts with NAC domains. Of these, 7 ZmNACs were putative targets for regulation by miR164. We isolated one gene, called TC258020 (designated ZmNAC1) from 2 maize inbred lines, 87-1 and Zong3. ZmNAC1 had a high expression level in roots and showed higher abundance (1.8 fold) in Zong3 relative to 87-1, which had less lateral roots than Zong3. There was a significant correlation between the expression level of ZmNAC1 and the lateral root density in the recombinant inbred line (RIL) population. Transgenic Arabidopsis that overexpressed ZmNAC1 had increased lateral roots in comparison to the wild type. These findings suggest that ZmNAC1 played a significant role in lateral root development. An allelic expression assay showed that trans-regulatory elements were the dominant mediators of ZmNAC1 differential expression in 87-1 and Zong3, and further analysis revealed that miR164 was a trans-element that guided the cleavage of endogenous ZmNAC1 mRNA. Both mature miR164 and miR164 precursors had higher expression in 87-1 than Zong3, which was the opposite of the expression pattern of ZmNAC1. Additionally, the allelic assay showed that the cis-regulatory element most likely affected Zm-miR164b's expression pattern. A β-glucuronidase (GUS) assay showed that the Zm-miR164b promoter had higher GUS activity in 87-1 than in Zong3. In addition, we detected miR164b expression in the RIL population, and the results indicated that miR164b had a higher expression level in the RILs containing 87-1 promoter than those containing Zong3 promoter.ConclusionOur results indicate one possible pathway in maize by which differences in miR164b promoter activity resulted in a different expression pattern for mature miR164 which negatively regulates ZmNAC1 expression in 87-1 and Zong3, thereby contributing to a significantly different lateral root phenotype.

BackgroundMicroRNAs (miRNAs) are a class of small non-coding regulatory RNAs that regulate gene expression by guiding target mRNA cleavage or translational inhibition. MiRNAs can have large-scale regulatory effects on development and stress response in plants.ResultsTo test whether miRNAs play roles in regulating response to powdery mildew infection and heat stress in wheat, by using Solexa high-throughput sequencing we cloned the small RNA from wheat leaves infected by preponderant physiological strain Erysiphe graminis f. sp. tritici (Egt) or by heat stress treatment. A total of 153 miRNAs were identified, which belong to 51 known and 81 novel miRNA families. We found that 24 and 12 miRNAs were responsive to powdery mildew infection and heat stress, respectively. We further predicted that 149 target genes were potentially regulated by the novel wheat miRNA.ConclusionsOur results indicated that diverse set of wheat miRNAs were responsive to powdery mildew infection and heat stress and could function in wheat responses to both biotic and abiotic stresses.

BackgroundHeat stress is one of the most crucial environmental factors, which reduces crop yield worldwide. In plants, the MYB family is one of the largest families of transcription factors (TFs). Although some wheat stress-related MYB TFs have been characterized, their involvement in response to high-temperature stress has not been properly studied.ResultsSix novel heat-induced MYB genes were identified by comparison with previously established de novo transcriptome sequencing data obtained from wheat plants subjected to heat treatment; genomic and complete coding sequences of these genes were isolated. All six TaMYBs were localized in the nucleus of wheat protoplasts. Transactivation assays in yeast revealed that all six proteins acted as transcriptional activators, and the activation domains were attributed to the C-termini of the six wheat MYB proteins. Phylogenetic analysis of the six TaMYBs and R2R3-MYBs from Arabidopsis revealed that all six proteins were in clades that contained stress-related MYB TFs. The expression profiles of TaMYB genes were different in wheat tissues and in response to various abiotic stresses and exogenous abscisic acid treatment. In transgenic Arabidopsis plants carrying TaMYB80 driven by the CaMV 35S promoter, tolerance to heat and drought stresses increased, which could be attributed to the increased levels of cellular abscisic acid.ConclusionsWe identified six heat-induced MYB genes in wheat. We performed comprehensive analyses of the cloned MYB genes and their gene products, including gene structures, subcellular localization, transcriptional activation, phylogenetic relationships, and expression patterns in different wheat tissues and under various abiotic stresses. In particular, we showed that TaMYB80 conferred heat and drought tolerance in transgenic Arabidopsis. These results contribute to our understanding of the functions of heat-induced MYB genes and provide the basis for selecting the best candidates for in-depth functional studies of heat-responsive MYB genes in wheat.

BackgroundThe yield of wheat (Triticum aestivum L.), an important crop, is adversely affected by heat stress in many regions of the world. However, the molecular mechanisms underlying thermotolerance are largely unknown.ResultsA novel ferritin gene, TaFER, was identified from our previous heat stress-responsive transcriptome analysis of a heat-tolerant wheat cultivar (TAM107). TaFER was mapped to chromosome 5B and named TaFER-5B. Expression pattern analysis revealed that TaFER-5B was induced by heat, polyethylene glycol (PEG), H2O2 and Fe-ethylenediaminedi(o-hydroxyphenylacetic) acid (Fe-EDDHA). To confirm the function of TaFER-5B in wheat, TaFER-5B was transformed into the wheat cultivar Jimai5265 (JM5265), and the transgenic plants exhibited enhanced thermotolerance. To examine whether the function of ferritin from mono- and dico-species is conserved, TaFER-5B was transformed into Arabidopsis, and overexpression of TaFER-5B functionally complemented the heat stress-sensitive phenotype of a ferritin-lacking mutant of Arabidopsis. Moreover, TaFER-5B is essential for protecting cells against heat stress associated with protecting cells against ROS. In addition, TaFER-5B overexpression also enhanced drought, oxidative and excess iron stress tolerance associated with the ROS scavenging. Finally, TaFER-5B transgenic Arabidopsis and wheat plants exhibited improved leaf iron content.ConclusionsOur results suggest that TaFER-5B plays an important role in enhancing tolerance to heat stress and other abiotic stresses associated with the ROS scavenging.