Detachment of plant organs occurs in abscission zones (AZs). During plant growth, the AZ forms, but does not develop further until the cells perceive abscission-promoting signals and initiate detachment. Upon signal perception, abscission initiates immediately; if there is no signal, abscission is not induced and the organ remains attached to the plant. However, little attention has been paid to the genes that maintain competence to respond to the abscission signal in the pre-abscission AZ. Recently, we found that the tomato (Solanum lycopersicum) transcription factors BLIND (Bl), GOBLET (GOB), Lateral suppressor (Ls) and a tomato WUSCHEL homologue (LeWUS) are expressed specifically in pre-abscission tissue, the anthesis pedicel AZs. To advance our understanding of abscission, here we profiled genome-wide gene expression in tomato flower pedicels at the pre-abscission stage.
We examined the transcriptomes of three tomato flower pedicel regions, the AZ and flanking proximal- (Prox) and distal- (Dis) regions, and identified 89 genes that were preferentially expressed in the AZ compared to both Prox and Dis. These genes included several transcription factors that regulate apical or axillary shoot meristem activity. Also, genes associated with auxin activity were regulated in a Prox-Dis region-specific manner, suggesting that a gradient of auxin exists in the pedicel. A MADS-box gene affecting floral transition was preferentially expressed in the Prox region and other MADS-box genes for floral organ identification were preferentially expressed in Dis, implying that the morphologically similar Prox and Dis regions have distinct identities. We also analyzed the expression of known regulators; in anthesis pedicels, Bl, GOB, Ls and LeWUS were expressed in the vascular cells of the AZ region. However, after an abscission signal, Bl was up-regulated, but GOB, Ls and LeWUS were down-regulated, suggesting that Bl may be a positive regulator of abscission, but the others may be negative regulators.
This study reveals region-specific gene expression in tomato flower pedicels at anthesis and identifies factors that may determine the physiological properties of the pre-abscission pedicel. The region-specific transcriptional regulators and genes for auxin activity identified here may prevent flower abscission in the absence of signal or establish competence to respond to the abscission signal.
Tomato (Solanum lycopersicum); Abscission zone; Flower pedicel; Auxin; Shoot meristem; Competence to respond to abscission signals; Transcription factor; MADS-box genes
Apple is a widely cultivated fruit crop for its quality properties and extended storability. Among the several quality factors, texture is the most important and appreciated, and within the apple variety panorama the cortex texture shows a broad range of variability. Anatomically these variations depend on degradation events occurring in both fruit primary cell wall and middle lamella. This physiological process is regulated by an enzymatic network generally encoded by large gene families, among which polygalacturonase is devoted to the depolymerization of pectin. In apple, Md-PG1, a key gene belonging to the polygalacturonase gene family, was mapped on chromosome 10 and co-localized within the statistical interval of a major hot spot QTL associated to several fruit texture sub-phenotypes.
In this work, a QTL corresponding to the position of Md-PG1 was validated and new functional alleles associated to the fruit texture properties in 77 apple cultivars were discovered. 38 SNPs genotyped by gene full length resequencing and 2 SSR markers ad hoc targeted in the gene metacontig were employed. Out of this SNP set, eleven were used to define three significant haplotypes statistically associated to several texture components. The impact of Md-PG1 in the fruit cell wall disassembly was further confirmed by the cortex structure electron microscope scanning in two apple varieties characterized by opposite texture performance, such as ‘Golden Delicious’ and ‘Granny Smith’.
The results here presented step forward into the genetic dissection of fruit texture in apple. This new set of haplotypes, and microsatellite alleles, can represent a valuable toolbox for a more efficient parental selection as well as the identification of new apple accessions distinguished by superior fruit quality features.
Over 200 published studies of more than 30 plant species have reported a role for miRNAs in regulating responses to abiotic stresses. However, data from these individual reports has not been collected into a single database. The lack of a curated database of stress-related miRNAs limits research in this field, and thus a cohesive database system should necessarily be constructed for data deposit and further application.
PASmiR, a literature-curated and web-accessible database, was developed to provide detailed, searchable descriptions of miRNA molecular regulation in different plant abiotic stresses. PASmiR currently includes data from ~200 published studies, representing 1038 regulatory relationships between 682 miRNAs and 35 abiotic stresses in 33 plant species. PASmiR’s interface allows users to retrieve miRNA-stress regulatory entries by keyword search using plant species, abiotic stress, and miRNA identifier. Each entry upon keyword query contains detailed regulation information for a specific miRNA, including species name, miRNA identifier, stress name, miRNA expression pattern, detection method for miRNA expression, a reference literature, and target gene(s) of the miRNA extracted from the corresponding reference or miRBase. Users can also contribute novel regulatory entries by using a web-based submission page. The PASmiR database is freely accessible from the two URLs of
The PASmiR database provides a solid platform for collection, standardization, and searching of miRNA-abiotic stress regulation data in plants. As such this database will be a comprehensive repository for miRNA regulatory mechanisms involved in plant response to abiotic stresses for the plant stress physiology community.
Previously, we have reported the ability of thiamine (vitamin B1) to induce resistance against Plasmopara viticola in a susceptible grapevine cv. Chardonnay. However, mechanisms underlying vitamins, especially, thiamine-induced disease resistance in grapevine are still largely unknown. Here, we assessed whether thiamine could modulate phenylpropanoid pathway-derived phytoalexins in grapevine plants, as well as, the role of such secondary metabolites in thiamine-induced resistance process to P. viticola.
Our data show that thiamine treatment elicited the expression of phenylpropanoid pathway genes in grapevine plants. The expression of these genes correlated with an accumulation of stilbenes, phenolic compounds, flavonoids and lignin. Furthermore, the total anti-oxidant potential of thiamine-treaded plants was increased by 3.5-fold higher level as compared with untreated-control plants. Four phenolic compounds are responsible of 97% of the total anti-oxidant potential of thiamine-treated plants. Among these compounds, is the caftaric acid, belonging to the hydroxy-cinnamic acids family. This element contributed, by its own, by 20% of this total anti-oxidant potential. Epifluorescence microscopy analysis revealed a concomitant presence of unbranched-altered P. viticola mycelia and stilbenes production in the leaf mesophyll of thiamine-treated inoculated plants, suggesting that stilbenes are an important component of thiamine-induced resistance in grapevine.
This work is the first to show the role of thiamine, as a vitamin, in the modulation of grapevine plant secondary metabolism contributing to an enhanced resistance to P. viticola, the most destructive fungal disease in vineyards.
Thiamine; Plasmopara viticola; Stilbenes; Phenylpropanoid pathway genes; Lignin; Flavonoids; Grapevine; Induced resistance; Real-Time q-PCR; HPLC; UPLC-MS
The selective removal of grapevine leaves around berry clusters can improve the quality of ripening fruits by influencing parameters such as the berry sugar and anthocyanin content at harvest. The outcome depends strongly on the timing of defoliation, which influences the source–sink balance and the modified microclimate surrounding the berries. We removed the basal leaves from Vitis vinifera L. cv Sangiovese shoots at the pre-bloom and veraison stages, and investigated responses such as shoot growth, fruit morphology and composition compared to untreated controls. Moreover, we performed a genome-wide expression analysis to explore the impact of these defoliation treatments on berry transcriptome.
We found that pre-bloom defoliation improved berry quality traits such as sugar and anthocyanin content, whereas defoliation at veraison had a detrimental effect, e.g. less anthocyanin and higher incidence of sunburn damage. Genome-wide expression analysis during berry ripening revealed that defoliation at either stage resulted in major transcriptome reprogramming, which slightly delayed the onset of ripening. However, a closer investigation of individual gene expression profiles identified genes that were specifically modulated by defoliation at each stage, reflecting the uncoupling of metabolic processes such as flavonoid biosynthesis, cell wall and stress metabolism, from the general ripening program.
The specific transcriptional modifications we observed following defoliation at different time points allow the identification of the developmental or metabolic processes affected in berries thus deepening the knowledge of the mechanisms by which these agronomical practices impact the final berry ripening traits.
Vitis vinifera; Defoliation; Berry ripening; Transcriptome; Flavonoid; Source-sink balance
Long-lived trees can accumulate mutations throughout their lifetimes that may influence biotic and abiotic interactions. For example, some Eucalyptus trees display marked variation in herbivore defence within a single canopy. These “mosaic” trees support foliage with distinct chemotypes which are differentially favoured by insect and vertebrate herbivores, resulting in susceptible and resistant branches within a single canopy. These mosaic trees provide a unique opportunity to explore the biosynthesis and genetic regulation of chemical defences in the foliage. The biosynthesis of the principal defence compounds, terpenoid-dominated essential oils, is well understood. However, the regulation of the genes involved and thus the control of phenotypic variation within a single tree canopy remains a mystery.
We sequenced the transcriptomes of the leaves of the two different chemotypes of a chemically mosaic Eucalyptus melliodora tree using 454 pyrosequencing technology. We used gene set enrichment analysis to identify differentially expressed transcripts and found the proportion of differentially expressed genes in the resistant and susceptible foliage similar to the transcript difference between functionally distinct tissues of the same organism, for example roots and leaves. We also investigated sequence differences in the form of single nucleotide polymorphisms and found 10 nucleotides that were different between the two branches. These are likely true SNPs and several occur in regulatory genes.
We found three lines of evidence that suggest changes to a ‘master switch’ can result in large scale phenotypic changes: 1. We found differential expression of terpene biosynthetic genes between the two chemotypes that could contribute to chemical variation within this plant. 2. We identified many genes that are differentially expressed between the two chemotypes, including some unique genes in each branch. These genes are involved in a variety of processes within the plant and many could contribute to the regulation of secondary metabolism, thus contributing to the chemical variation. 3. We identified 10 SNPs, some of which occur in regulatory genes that could influence secondary metabolism and thus contribute to chemical variation. Whilst this research is inherently limited by sample size, the patterns we describe could be indicative of other plant genetic mosaics.
Eucalyptus; Mosaic; Somatic mutation; Comparative transcriptomics; Herbivory
The editors of BMC Plant Biology would like to thank all of our reviewers who have contributed to the journal in Volume 12 (2012).
Plant parasitism represents an extraordinary interaction among flowering plants: parasitic plants use a specialized organ, the haustorium, to invade the host vascular system to deprive host plants of water and nutrients. Various compounds present in exudates of host plants trigger haustorium development. The two most effective haustorium inducing factors (HIFs) known for the parasitic plant Triphysaria versicolor (T. versicolor) are peonidin, an antioxidant flavonoid, and 2,6-dimethoxybenzoquinone (DMBQ), an oxidative stress agent. To date, two genes involved in haustorium initiation in T. versicolor have been identified: TvQR1, a quinone oxidoreductase that generates the active HIF from DMBQ, and TvPirin, a transcription co-factor that regulates several other DMBQ- responsive and –non-responsive genes. While the expression of these genes in response to DMBQ is well characterized, their expression in response to peonidin is not. In addition, the pattern of polymorphisms in these genes is unknown, even though nucleotide changes in TvQR1 and TvPirin may have contributed to the ability of T. versicolor to develop haustoria. To gain insights into these aspects, we investigated their transcriptional responses to HIFs and non-HIF and their natural nucleotide diversity.
Here we show that TvQR1 and TvPirin are transcriptionally upregulated by both DMBQ and peonidin in T. versicolor roots. Yet, while TvQR1 also responded to juglone, a non-HIF quinone with toxicity comparable to that of DMBQ, TvPirin did not. We further demonstrate that TvPirin encodes a protein shorter than the one previously reported. In the T. versicolor natural population of Northern California, TvQR1 exhibited remarkably higher molecular diversity and more recombination events than TvPirin, with the highest non-synonymous substitution rate in the substrate recognition and catalytic domain of the TvQR1 protein.
Our results suggest that TvQR1 and TvPirin have most likely evolved highly distinct roles for haustorium formation. Unlike TvPirin, TvQR1 might have been under diversifying selection to maintain a diverse collection of polymorphisms, which might be related to the recognition of an assortment of HIF and non-HIF quinones as substrates for successful haustorial establishment in a wide range of host plants.
Parasitic plants; Haustorium; Nucleotide diversity; Allelic polymorphism; Population genetics; Transcriptional responses
Grapevine (Vitis vinifera subsp. vinifera) is one of the most important and ancient horticultural plants in the world. Domesticated about 8–10,000 years ago in the Eurasian region, grapevine evolved from its wild relative (V. vinifera subsp. sylvestris) into very diverse and heterozygous cultivated forms. In this work we study grapevine genetic structure in a large sample of cultivated varieties, to interpret the wide diversity at morphological and molecular levels and link it to cultivars utilization, putative geographic origin and historical events.
We analyzed the genetic structure of cultivated grapevine using a dataset of 2,096 multi-locus genotypes defined by 20 microsatellite markers. We used the Bayesian approach implemented in the STRUCTURE program and a hierarchical clustering procedure based on Ward’s method to assign individuals to sub-groups. The analysis revealed three main genetic groups defined by human use and geographic origin: a) wine cultivars from western regions, b) wine cultivars from the Balkans and East Europe, and c) a group mainly composed of table grape cultivars from Eastern Mediterranean, Caucasus, Middle and Far East countries. A second structure level revealed two additional groups, a geographic group from the Iberian Peninsula and Maghreb, and a group comprising table grapes of recent origins from Italy and Central Europe. A large number of admixed genotypes were also identified. Structure clusters regrouped together a large proportion of family-related genotypes. In addition, Ward’s method revealed a third level of structure, corresponding either to limited geographic areas, to particular grape use or to family groups created through artificial selection and breeding.
This study provides evidence that the cultivated compartment of Vitis vinifera L. is genetically structured. Genetic relatedness of cultivars has been shaped mostly by human uses, in combination with a geographical effect. The finding of a large portion of admixed genotypes may be the trace of both large human-mediated exchanges between grape-growing regions throughout history and recent breeding.
SKIP is a transcription cofactor in many eukaryotes. It can regulate plant stress tolerance in rice and Arabidopsis. But the homolog of SKIP protein in soybean has been not reported up to now.
In this study, the expression patterns of soybean GAMYB binding protein gene (GmGBP1) encoding a homolog of SKIP protein were analyzed in soybean under abiotic stresses and different day lengths. The expression of GmGBP1 was induced by polyethyleneglycol 6000, NaCl, gibberellin, abscisic acid and heat stress. GmGBP1 had transcriptional activity in C-terminal. GmGBP1 could interact with R2R3 domain of GmGAMYB1 in SKIP domain to take part in gibberellin flowering pathway. In long-day (16 h-light) condition, transgenic Arabidopsis with the ectopic overexpression of GmGBP1 exhibited earlier flowering and less number of rosette leaves; Suppression of AtSKIP in Arabidopsis resulted in growth arrest, flowering delay and down-regulation of many flowering-related genes (CONSTANS, FLOWERING LOCUS T, LEAFY); Arabidopsis myb33 mutant plants with ectopic overexpression of GmGBP1 showed the same flowering phenotype with wild type. In short-day (8 h-light) condition, transgenic Arabidopsis plants with GmGBP1 flowered later and showed a higher level of FLOWERING LOCUS C compared with wild type. When treated with abiotic stresses, transgenic Arabidopsis with the ectopic overexpression of GmGBP1 enhanced the tolerances to heat and drought stresses but reduced the tolerance to high salinity, and affected the expressions of several stress-related genes.
In Arabidopsis, GmGBP1 might positively regulate the flowering time by affecting CONSTANS, FLOWERING LOCUS T, LEAFY and GAMYB directly or indirectly in photoperiodic and gibberellin pathways in LDs, but GmGBP1 might represse flowering by affecting FLOWERING LOCUS C and SHORT VEGETATIVE PHASE in autonomous pathway in SDs. GmGBP1 might regulate the activity of ROS-eliminating to improve the resistance to heat and drought but reduce the high-salinity tolerance.
GmGBP1; Abiotic stress; Flowering; Day-length; Gibberellin
Abundance and distribution of the plant hormone auxin play important roles in plant development. Besides other metabolic processes, various auxin carriers control the cellular level of active auxin and, hence, are major regulators of cellular auxin homeostasis. Despite the developmental importance of auxin transporters, a simple medium-to-high throughput approach to assess carrier activities is still missing. Here we show that carrier driven depletion of cellular auxin correlates with reduced nuclear auxin signaling in tobacco Bright Yellow-2 (BY-2) cell cultures.
We developed an easy to use transient single-cell-based system to detect carrier activity. We use the relative changes in signaling output of the auxin responsive promoter element DR5 to indirectly visualize auxin carrier activity. The feasibility of the transient approach was demonstrated by pharmacological and genetic interference with auxin signaling and transport. As a proof of concept, we provide visual evidence that the prominent auxin transport proteins PIN-FORMED (PIN)2 and PIN5 regulate cellular auxin homeostasis at the plasma membrane and endoplasmic reticulum (ER), respectively. Our data suggest that PIN2 and PIN5 have different sensitivities to the auxin transport inhibitor 1-naphthylphthalamic acid (NPA). Also the putative PIN-LIKES (PILS) auxin carrier activity at the ER is insensitive to NPA in our system, indicating that NPA blocks intercellular, but not intracellular auxin transport.
This single-cell-based system is a useful tool by which the activity of putative auxin carriers, such as PINs, PILS and WALLS ARE THIN1 (WAT1), can be indirectly visualized in a medium-to-high throughput manner. Moreover, our single cell system might be useful to investigate also other hormonal signaling pathways, such as cytokinin.
Auxin homeostasis; DR5; Auxin carrier; Auxin transport
During plant tissue morphogenesis cells have to coordinate their behavior to allow the generation of the size, shape and cellular patterns that distinguish an organ. Despite impressive progress the underlying signaling pathways remain largely unexplored. In Arabidopsis thaliana, the atypical leucine-rich repeat receptor-like kinase STRUBBELIG (SUB) is involved in signal transduction in several developmental processes including the formation of carpels, petals, ovules and root hair patterning. The three STRUBBELIG-LIKE MUTANT (SLM) genes DETORQUEO (DOQ), QUIRKY (QKY) and ZERZAUST (ZET) are considered central elements of SUB-mediated signal transduction pathways as corresponding mutants share most phenotypic aspects with sub mutants.
Here we show that DOQ corresponds to the previously identified ANGUSTIFOLIA gene. The genetic analysis revealed that the doq-1 mutant exhibits all additional mutant phenotypes and conversely that other an alleles show the slm phenotypes. We further provide evidence that SUB and AN physically interact and that AN is not required for subcellular localization of SUB.
Our data suggest that AN is involved in SUB signal transduction pathways. In addition, they reveal previously unreported functions of AN in several biological processes, such as ovule development, cell morphogenesis in floral meristems, and root hair patterning. Finally, SUB and AN may directly interact at the plasma membrane to mediate SUB-dependent signaling.
ANGUSTIFOLIA; Flower; Ovule; Receptor-like kinase; Root hair patterning; Signal transduction; Tissue morphogenesis; STRUBBELIG
Maize (Zea mays L.) leaves damaged by lepidopteran herbivores emit a complex volatile blend that can attract natural enemies of the herbivores and may also have roles in direct defense and inter- or intra-plant signaling. The volatile blend is dominated by sesquiterpenes of which the majority is produced by two herbivore-induced terpene synthases, TPS10 and TPS23. However, little is known about the pattern of volatile emission within maize leaves.
In this study, we restricted herbivore feeding to small sections of the maize leaf with the aim of determining the patterns of volatile sesquiterpene emission throughout the damaged leaf and in neighboring leaves. Sesquiterpene volatiles were released at high rates from damaged leaves, but at much lower rates from neighboring leaves. Release was restricted to the site of damage or to leaf sections located apical to the damage, but was not seen in sections basal to the damage or on the other side of the midrib. The emission pattern correlated well with the transcript pattern of the respective sesquiterpene synthase genes, tps10 and tps23, implying that biosynthesis likely occurs at the site of emission. The concentrations of jasmonic acid and its leucine derivative were also elevated in terpene-emitting tissues suggesting a role for jasmonates in propagating the damage signal.
In contrast to other defense reactions which often occur systemically throughout the whole plant, herbivore-induced sesquiterpene production in maize is restricted to the wounding site and distal leaf parts. Since the signal mediating this reaction is directed to the leaf tip and cannot propagate parallel to the leaf axis, it is likely connected to the xylem. The increasing gradient of volatiles from the tip of the leaf towards the damage site might aid herbivore enemies in host or prey finding.
Zea mays L.; Poaceae; Maize; Sesquiterpenes; Volatiles; Terpene biosynthesis; Terpene synthase; Herbivore-induced terpene formation; Jasmonic acid; Solid-phase microextraction (SPME)
Bitter acids (e.g. humulone) are prenylated polyketides synthesized in lupulin glands of the hop plant (Humulus lupulus) which are important contributors to the bitter flavour and stability of beer. Bitter acids are formed from acyl-CoA precursors derived from branched-chain amino acid (BCAA) degradation and C5 prenyl diphosphates from the methyl-D-erythritol 4-phosphate (MEP) pathway. We used RNA sequencing (RNA-seq) to obtain the transcriptomes of isolated lupulin glands, cones with glands removed and leaves from high α-acid hop cultivars, and analyzed these datasets for genes involved in bitter acid biosynthesis including the supply of major precursors. We also measured the levels of BCAAs, acyl-CoA intermediates, and bitter acids in glands, cones and leaves.
Transcripts encoding all the enzymes of BCAA metabolism were significantly more abundant in lupulin glands, indicating that BCAA biosynthesis and subsequent degradation occurs in these specialized cells. Branched-chain acyl-CoAs and bitter acids were present at higher levels in glands compared with leaves and cones. RNA-seq analysis showed the gland-specific expression of the MEP pathway, enzymes of sucrose degradation and several transcription factors that may regulate bitter acid biosynthesis in glands. Two branched-chain aminotransferase (BCAT) enzymes, HlBCAT1 and HlBCAT2, were abundant, with gene expression quantification by RNA-seq and qRT-PCR indicating that HlBCAT1 was specific to glands while HlBCAT2 was present in glands, cones and leaves. Recombinant HlBCAT1 and HlBCAT2 catalyzed forward (biosynthetic) and reverse (catabolic) reactions with similar kinetic parameters. HlBCAT1 is targeted to mitochondria where it likely plays a role in BCAA catabolism. HlBCAT2 is a plastidial enzyme likely involved in BCAA biosynthesis. Phylogenetic analysis of the hop BCATs and those from other plants showed that they group into distinct biosynthetic (plastidial) and catabolic (mitochondrial) clades.
Our analysis of the hop transcriptome significantly expands the genomic resources available for this agriculturally-important crop. This study provides evidence for the lupulin gland-specific biosynthesis of BCAAs and prenyl diphosphates to provide precursors for the production of bitter acids. The biosynthetic pathway leading to BCAAs in lupulin glands involves the plastidial enzyme, HlBCAT2. The mitochondrial enzyme HlBCAT1 degrades BCAAs as the first step in the catabolic pathway leading to branched chain-acyl-CoAs.
Lettuce (Lactuca sativa L.) is the major crop from the group of leafy vegetables. Several types of molecular markers were developed that are effectively used in lettuce breeding and genetic studies. However only a very limited number of microsattelite-based markers are publicly available. We have employed the method of enriched microsatellite libraries to develop 97 genomic SSR markers.
Testing of newly developed markers on a set of 36 Lactuca accession (33 L. sativa, and one of each L. serriola L., L. saligna L., and L. virosa L.) revealed that both the genetic heterozygosity (UHe = 0.56) and the number of loci per SSR (Na = 5.50) are significantly higher for genomic SSR markers than for previously developed EST-based SSR markers (UHe = 0.32, Na = 3.56). Fifty-four genomic SSR markers were placed on the molecular linkage map of lettuce. Distribution of markers in the genome appeared to be random, with the exception of possible cluster on linkage group 6. Any combination of 32 genomic SSRs was able to distinguish genotypes of all 36 accessions. Fourteen of newly developed SSR markers originate from fragments with high sequence similarity to resistance gene candidates (RGCs) and RGC pseudogenes. Analysis of molecular variance (AMOVA) of L. sativa accessions showed that approximately 3% of genetic diversity was within accessions, 79% among accessions, and 18% among horticultural types.
The newly developed genomic SSR markers were added to the pool of previously developed EST-SSRs markers. These two types of SSR-based markers provide useful tools for lettuce cultivar fingerprinting, development of integrated molecular linkage maps, and mapping of genes.
Data resolution statistics; Genotyping; Lactuca; Linkage map; Marker distribution; Microsatellites
Alternate bearing is a widespread phenomenon among crop plants, defined as the tendency of certain fruit trees to produce a high-yield crop one year ("on-year"), followed by a low-yield or even no crop the following year ("off-year"). Several factors may affect the balance between such developmental phase-transition processes. Among them are the microRNA (miRNA), being gene-expression regulators that have been found to be involved as key determinants in several physiological processes.
Six olive (Olea europaea L. cv. Ayvalik variety) small RNA libraries were constructed from fruits (ripe and unripe) and leaves (”on year” and ”off year” leaves in July and in November, respectively) and sequenced by high-throughput Illumina sequencing. The RNA was retrotranscribed and sequenced using the high-throughput Illumina platform. Bioinformatics analyses of 93,526,915 reads identified 135 conserved miRNA, belonging to 22 miRNA families in the olive. In addition, 38 putative novel miRNAs were discovered in the datasets. Expression of olive tree miRNAs varied greatly among the six libraries, indicating the contribution of diverse miRNA in balancing between reproductive and vegetative phases. Predicted targets of miRNA were categorized into 108 process ontology groups with significance abundance. Among those, potential alternate bearing-associated processes were found, such as development, hormone-mediated signaling and organ morphogenesis. The KEGG analyses revealed that the miRNA-targeted genes are involved in seven main pathways, belonging to carbohydrate metabolism and hormone signal-transduction pathways.
A comprehensive study on olive miRNA related to alternate bearing was performed. Regulation of miRNA under different developmental phases and tissues indicated that control of nutrition and hormone, together with flowering processes had a noteworthy impact on the olive tree alternate bearing. Our results also provide significant data on the miRNA-fruit development interaction and advance perspectives in the miRNA profile of the olive tree.
High-throughput small RNA sequencing; MicroRNA; Olive; Periodicity
Arabinogalactan proteins (AGPs) are cell wall proteoglycans that have been shown to be important for pollen development. An Arabidopsis double null mutant for two pollen-specific AGPs (agp6 agp11) showed reduced pollen tube growth and compromised response to germination cues in vivo. A microarray experiment was performed on agp6 agp11 pollen tubes to search for genetic interactions in the context of pollen tube growth. A yeast two-hybrid experiment for AGP6 and AGP11 was also designed.
The lack of two specific AGPs induced a meaningful shift in the gene expression profile. In fact, a high number of genes showed altered expression levels, strengthening the case that AGP6 and AGP11 are involved in complex phenomena. The expression levels of calcium- and signaling-related genes were found to be altered, supporting the known roles of the respective proteins in pollen tube growth. Although the precise nature of the proposed interactions needs further investigation, the putative involvement of AGPs in signaling cascades through calmodulin and protein degradation via ubiquitin was indicated. The expression of stress-, as well as signaling- related, genes was also changed; a correlation that may result from the recognized similarities between signaling pathways in both defense and pollen tube growth.
The results of yeast two-hybrid experiments lent further support to these signaling pathways and revealed putative AGP6 and AGP11 interactors implicated in recycling of cell membrane components via endocytosis, through clathrin-mediated endosomes and multivesicular bodies.
The data presented suggest the involvement of AGP6 and AGP11 in multiple signaling pathways, in particular those involved in developmental processes such as endocytosis-mediated plasma membrane remodeling during Arabidopsis pollen development. This highlights the importance of endosomal trafficking pathways which are rapidly emerging as fundamental regulators of the wall physiology.
Arabidopsis; Arabinogalactan proteins; Pollen tube; Microarray; Yeast two-hybrid
RNA silencing affects a broad range of regulatory processes in all eukaryotes ranging from chromatin structure maintenance to transcriptional and translational regulation and longevity of the mRNAs. Particularly in plants, it functions as the major defense mechanism against viruses. To counter-act this defense, plant viruses produce suppressors of RNA silencing (Viral suppressors of RNA silencing, VSRSs), which are essential for viruses to invade their specific host plants. Interactions of these VSRSs with the hosts’ silencing pathways, and their direct and indirect interference with different cellular regulatory networks constitute one of the main lines of the molecular virus-host interactions. Here we have used a microarray approach to study the effects of the Potato virus X Potexvirus (PVX)-specific P25 VSRS protein on the transcript profile of tobacco plants, when expressed as a transgene in these plants.
The expression of the PVX-specific P25 silencing suppressor in transgenic tobacco plants caused significant up-regulation of 1350 transcripts, but down-regulation of only five transcripts in the leaves, and up- and down-regulation of 51 and 13 transcripts, respectively, in the flowers of these plants, as compared to the wild type control plants. Most of the changes occurred in the transcripts related to biotic and abiotic stresses, transcription regulation, signaling, metabolic pathways and cell wall modifications, and many of them appeared to be induced through up-regulation of the signaling pathways regulated by ethylene, jasmonic acid and salicylic acid. Correlations of these alterations with the protein profile and related biological functions were analyzed. Surprisingly, they did not cause significant alterations in the protein profile, and caused only very mild alteration in the phenotype of the P25-expressing transgenic plants.
Expression of the PVX-specific P25 VSRS protein causes major alterations in the transcriptome of the leaves of transgenic tobacco plants, but very little of any effects in the young flowers of the same plants. The fairly stable protein profile in the leaves and lack of any major changes in the plant phenotype indicate that the complicated interplay and interactions between different regulatory levels are able to maintain homeostasis in the plants.
The transitions from juvenile to adult and adult to reproductive phases of growth are important stages in the life cycle of plants. The regulators of these transitions include miRNAs, in particular miR156 and miR172 which are part of a regulatory module conserved across the angiosperms. In Arabidopsis miR171 represses differentiation of axillary meristems by repressing expression of SCARECROW-LIKE(SCL) transcription factors, however the role of miR171 has not been examined in other plants.
To investigate the roles of mir171 and its target genes in a monocot, the Hvu pri-miR171a was over-expressed in barley (Hordeum vulgare L. cv. Golden promise) leading to reduced expression of at least one HvSCL gene. The resulting transgenic plants displayed a pleiotropic phenotype which included branching defects, an increased number of short vegetative phytomers and late flowering. These phenotypes appear to be the consequence of changes in the organisation of the shoot meristem. In addition, the data show that miR171 over-expression alters the vegetative to reproductive phase transition by activating the miR156 pathway and repressing the expression of the TRD (THIRD OUTER GLUME) and HvPLA1 (Plastochron1) genes.
Our data suggest that some of the roles of miR171 and its target genes that have been determined in Arabidopsis are conserved in barley and that they have additional functions in barley including activation of the miR156 pathway.
Barley; miR171; Scarecrow-like; Phase change; Meristems; Flowering time
The application and nutritional value of vegetable oil is highly dependent on its fatty acid composition, especially the relative proportion of its two major fatty acids, i.e oleic acid and linoleic acid. Microsomal oleoyl phosphatidylcholine desaturase encoded by FAD2 gene is known to introduce a double bond at the Δ12 position of an oleic acid on phosphatidylcholine and convert it to linoleic acid. The known plant FAD2 enzymes are encoded by small gene families consisting of 1-4 members. In addition to the classic oleate Δ12-desaturation activity, functional variants of FAD2 that are capable of undertaking additional or alternative acyl modifications have also been reported in a limited number of plant species. In this study, our objective was to identify FAD2 genes from safflower and analyse their differential expression profile and potentially diversified functionality.
We report here the characterization and functional expression of an exceptionally large FAD2 gene family from safflower, and the temporal and spatial expression profiles of these genes as revealed through Real-Time quantitative PCR. The diversified functionalities of some of the safflower FAD2 gene family members were demonstrated by ectopic expression in yeast and transient expression in Nicotiana benthamiana leaves. CtFAD2-1 and CtFAD2-10 were demonstrated to be oleate desaturases specifically expressed in developing seeds and flower head, respectively, while CtFAD2-2 appears to have relatively low oleate desaturation activity throughout the plant. CtFAD2-5 and CtFAD2-8 are specifically expressed in root tissues, while CtFAD2-3, 4, 6, 7 are mostly expressed in the cotyledons and hypocotyls in young safflower seedlings. CtFAD2-9 was found to encode a novel desaturase operating on C16:1 substrate. CtFAD2-11 is a tri-functional enzyme able to introduce a carbon double bond in either cis or trans configuration, or a carbon triple (acetylenic) bond at the Δ12 position.
In this study, we isolated an unusually large FAD2 gene family with 11 members from safflower. The seed expressed FAD2 oleate Δ12 desaturase genes identified in this study will provide candidate targets to manipulate the oleic acid level in safflower seed oil. Further, the divergent FAD2 enzymes with novel functionality could be used to produce rare fatty acids, such as crepenynic acid, in genetically engineered crop plants that are precursors for economically important phytoalexins and oleochemical products.
Plant cell walls are complex multicomponent structures that have evolved to fulfil an essential function in providing strength and protection to cells. Hemicelluloses constitute a key component of the cell wall and recently a number of the genes thought to encode the enzymes required for its synthesis have been identified in Arabidopsis. The acquisition of hemicellulose synthesis capability is hypothesised to have been an important step in the evolution of higher plants.
Analysis of the Physcomitrella patens genome has revealed the presence of homologs for all of the Arabidopsis glycosyltransferases including IRX9, IRX10 and IRX14 required for the synthesis of the glucuronoxylan backbone. The Physcomitrella IRX10 homolog is expressed in a variety of moss tissues which were newly formed or undergoing expansion. There is a high degree of sequence conservation between the Physcomitrella IRX10 and Arabidopsis IRX10 and IRX10-L. Despite this sequence similarity, the Physcomitrella IRX10 gene is only able to partially rescue the Arabidopsis irx10 irx10-L double mutant indicating that there has been a neo- or sub-functionalisation during the evolution of higher plants. Analysis of the monosaccharide composition of stems from the partially rescued Arabidopsis plants does not show any significant change in xylose content compared to the irx10 irx10-L double mutant. Likewise, knockout mutants of the Physcomitrella IRX10 gene do not result in any visible phenotype and there is no significant change in monosaccharide composition of the cell walls.
The fact that the Physcomitrella IRX10 (PpGT47A) protein can partially complement an Arabidopsis irx10 irx10-L double mutant suggests that it shares some function with the Arabidopsis proteins, but the lack of a phenotype in knockout lines shows that the function is not required for growth or development under normal conditions in Physcomitrella. In contrast, the Arabidopsis irx10 and irx10 irx10-L mutants have strong phenotypes indicating an important function in growth and development. We conclude that the evolution of vascular plants has been associated with a significant change or adaptation in the function of the IRX10 gene family.
Plant cell wall; Hemicellulose; Arabidopsis thaliana; Physcomitrella patens; Glycosyltransferases; Xylan
The coordination of growth within a tissue layer is of critical importance for tissue morphogenesis. For example, cells within the epidermis undergo stereotypic cell divisions that are oriented along the plane of the layer (planar growth), thereby propagating the layered epidermal structure. Little is known about the developmental control that regulates such planar growth in plants. Recent evidence suggested that the Arabidopsis AGC VIII protein kinase UNICORN (UCN) maintains planar growth by suppressing the formation of ectopic multicellular protrusions in several floral tissues including integuments. In the current model UCN controls this process during integument development by directly interacting with the ABERRANT TESTA SHAPE (ATS) protein, a member of the KANADI (KAN) family of transcription factors, thereby repressing its activity. Here we report on the further characterization of the UCN mechanism.
Phenotypic analysis of flowers of ucn-1 plants impaired in floral homeotic gene activity revealed that any of the four floral whorls could produce organs carrying ucn-1 protrusions. The ectopic outgrowths of ucn integuments did not accumulate detectable signals of the auxin and cytokinin reporters DR5rev::GFP and ARR5::GUS, respectively. Furthermore, wild-type and ucn-1 seedlings showed similarly strong callus formation upon in vitro culture on callus-inducing medium. We also show that ovules of ucn-1 plants carrying the dominant ats allele sk21-D exhibited more pronounced protrusion formation. Finally ovules of ucn-1 ett-1 double mutants and ucn-1 ett-1 arf4-1 triple mutants displayed an additive phenotype.
These data deepen the molecular insight into the UCN-mediated control of planar growth during integument development. The presented evidence indicates that UCN downstream signaling does not involve the control of auxin or cytokinin homeostasis. The results also reveal that UCN interacts with ATS independently of an ATS/ETT complex required for integument initiation and they further emphasize the necessity to balance UCN and ATS proteins during maintenance of planar growth in integuments.
Arabidopsis; Auxin; ABERRANT TESTA SHAPE; AGC protein kinase; AUXIN RESPONSE FACTOR 4; Cell division; Cytokinin; ETTIN; Growth regulation; KANADI; Planar growth; Ovule; Signal transduction; UNICORN
The Arabidopsis thaliana gene SPATULA (SPT), encoding a bHLH transcription factor, was originally identified for its role in pistil development. SPT is necessary for the growth and development of all carpel margin tissues including the style, stigma, septum and transmitting tract. Since then, it has been shown to have pleiotropic roles during development, including restricting the meristematic region of the leaf primordia and cotyledon expansion. Although SPT is expressed in roots, its role in this organ has not been investigated.
An analysis of embryo and root development showed that loss of SPT function causes an increase in quiescent center size in both the embryonic and postembryonic stem cell niches. In addition, root meristem size is larger due to increased division, which leads to a longer primary root. spt mutants exhibit other pleiotropic developmental phenotypes, including more flowers, shorter internodes and an extended flowering period. Genetic and molecular analysis suggests that SPT regulates cell proliferation in parallel to gibberellic acid as well as affecting auxin accumulation or transport.
Our data suggest that SPT functions in growth control throughout sporophytic growth of Arabidopsis, but is not necessary for cell fate decisions except during carpel development. SPT functions independently of gibberellic acid during root development, but may play a role in regulating auxin transport or accumulation. Our data suggests that SPT plays a role in control of root growth, similar to its roles in above ground tissues.
Maize is a major crop plant, grown for human and animal nutrition, as well as a renewable resource for bioenergy. When looking at the problems of limited fossil fuels, the growth of the world’s population or the world’s climate change, it is important to find ways to increase the yield and biomass of maize and to study how it reacts to specific abiotic and biotic stress situations. Within the OPTIMAS systems biology project maize plants were grown under a large set of controlled stress conditions, phenotypically characterised and plant material was harvested to analyse the effect of specific environmental conditions or developmental stages. Transcriptomic, metabolomic, ionomic and proteomic parameters were measured from the same plant material allowing the comparison of results across different omics domains. A data warehouse was developed to store experimental data as well as analysis results of the performed experiments.
The OPTIMAS Data Warehouse (OPTIMAS-DW) is a comprehensive data collection for maize and integrates data from different data domains such as transcriptomics, metabolomics, ionomics, proteomics and phenomics. Within the OPTIMAS project, a 44K oligo chip was designed and annotated to describe the functions of the selected unigenes. Several treatment- and plant growth stage experiments were performed and measured data were filled into data templates and imported into the data warehouse by a Java based import tool. A web interface allows users to browse through all stored experiment data in OPTIMAS-DW including all data domains. Furthermore, the user can filter the data to extract information of particular interest. All data can be exported into different file formats for further data analysis and visualisation. The data analysis integrates data from different data domains and enables the user to find answers to different systems biology questions. Finally, maize specific pathway information is provided.
With OPTIMAS-DW a data warehouse for maize was established, which is able to handle different data domains, comprises several analysis results that will support researchers within their work and supports systems biological research in particular. The system is available at
Maize; Zea mays; Database; WGCNA; Biomass; Yield; Data integration; Transcriptomics; Metabolomics; Phenomics
Ethylene production and signalling play an important role in somatic embryogenesis, especially for species that are recalcitrant in in vitro culture. The AP2/ERF superfamily has been identified and classified in Hevea brasiliensis. This superfamily includes the ERFs involved in response to ethylene. The relative transcript abundance of ethylene biosynthesis genes and of AP2/ERF genes was analysed during somatic embryogenesis for callus lines with different regeneration potential, in order to identify genes regulated during that process.
The analysis of relative transcript abundance was carried out by real-time RT-PCR for 142 genes. The transcripts of ERFs from group I, VII and VIII were abundant at all stages of the somatic embryogenesis process. Forty genetic expression markers for callus regeneration capacity were identified. Fourteen markers were found for proliferating calli and 35 markers for calli at the end of the embryogenesis induction phase. Sixteen markers discriminated between normal and abnormal embryos and, lastly, there were 36 markers of conversion into plantlets. A phylogenetic analysis comparing the sequences of the AP2 domains of Hevea and Arabidopsis genes enabled us to predict the function of 13 expression marker genes.
This first characterization of the AP2/ERF superfamily in Hevea revealed dramatic regulation of the expression of AP2/ERF genes during the somatic embryogenesis process. The gene expression markers of proliferating callus capacity to regenerate plants by somatic embryogenesis should make it possible to predict callus lines suitable to be used for multiplication. Further functional characterization of these markers opens up prospects for discovering specific AP2/ERF functions in the Hevea species for which somatic embryogenesis is difficult.
Gene expression; Plant hormone; Plant regeneration; Recalcitrant; Rubber; Signalling; Transcription factor