Songtao Jiu

Abscisic acid and sucrose regulate tomato and strawberry fruit ripening through the abscisic acid-stress-ripening transcription factor.

Summary Although great progress has been made towards understanding the role of abscisic acid (ABA) and sucrose in fruit ripening, the mechanisms underlying the ABA and sucrose signalling pathways remain elusive. In this study, transcription factor ABA‐stress‐ripening (ASR), which is involved in the transduction of ABA and sucrose signalling pathways, was isolated and analysed in the nonclimacteric fruit, strawberry and the climacteric fruit, tomato. We have identified four ASR isoforms in tomato and one in strawberry. All ASR sequences contained the ABA stress‐ and ripening‐induced proteins and water‐deficit stress‐induced proteins (ABA/WDS) domain and all ASR transcripts showed increased expression during fruit development. The expression of the ASR gene was influenced not only by sucrose and ABA, but also by jasmonic acid (JA) and indole‐3‐acetic acid (IAA), and these four factors were correlated with each other during fruit development. ASR bound the hexose transporter (HT) promoter, which contained a sugar box that activated downstream gene expression. Overexpression of the ASR gene promoted fruit softening and ripening, whereas RNA interference delayed fruit ripening, as well as affected fruit physiological changes. Change in ASR gene expression influenced the expression of several ripening‐related genes such as CHS,CHI, F3H,DFR,ANS,UFGT,PG,PL,EXP1/2,XET16, Cel1/2 and PME. Taken together, this study may provide new evidence on the important role of ASR in cross‐signalling between ABA and sucrose to regulate tomato and strawberry fruit ripening. The findings of this study also provide new insights into the regulatory mechanism underlying fruit development.

Genome-Wide Analysis of the Sucrose Synthase Gene Family in Grape (Vitis vinifera): Structure, Evolution, and Expression Profiles

Sucrose synthase (SS) is widely considered as the key enzyme involved in the plant sugar metabolism that is critical to plant growth and development, especially quality of the fruit. The members of SS gene family have been identified and characterized in multiple plant genomes. However, detailed information about this gene family is lacking in grapevine (Vitis vinifera L.). In this study, we performed a systematic analysis of the grape (V. vinifera) genome and reported that there are five SS genes (VvSS1–5) in the grape genome. Comparison of the structures of grape SS genes showed high structural conservation of grape SS genes, resulting from the selection pressures during the evolutionary process. The segmental duplication of grape SS genes contributed to this gene family expansion. The syntenic analyses between grape and soybean (Glycine max) demonstrated that these genes located in corresponding syntenic blocks arose before the divergence of grape and soybean. Phylogenetic analysis revealed distinct evolutionary paths for the grape SS genes. VvSS1/VvSS5, VvSS2/VvSS3 and VvSS4 originated from three ancient SS genes, which were generated by duplication events before the split of monocots and eudicots. Bioinformatics analysis of publicly available microarray data, which was validated by quantitative real-time reverse transcription PCR (qRT-PCR), revealed distinct temporal and spatial expression patterns of VvSS genes in various tissues, organs and developmental stages, as well as in response to biotic and abiotic stresses. Taken together, our results will be beneficial for further investigations into the functions of SS gene in the processes of grape resistance to environmental stresses.

Genome-Wide Mapping and Analysis of Grapevine MicroRNAs and Their Potential Target Genes

MicroRNAs (miRNAs) are single‐stranded, nonprotein‐coding, endogenously expressed, small RNAs 19 to 25 nucleotides in length. Recognizing the lack of specific and systematic studies on genome‐wide mapping of grapevine (Vitis vinifera L.) miRNAs, we conducted genome‐wide mapping of Vv‐miRNAs (V. vinifera miRNAs), SB‐miRNAs (V. vinifera L. ‘Summer Black’ miRNAs), and Va‐miRNAs (V. amurensis Rupr. miRNAs). The mapping results revealed that many of miRNAs located within the intergenic region had independent transcription units. To further validate the mapping results and existence of miRNAs, 12 randomly selected precursors of miRNAs (pre‐miRNAs) were successfully cloned and sequenced. Subsequently, 15 conserved and 29 nonconserved intragenic (intronic, exonic) Vv‐miRNA genes, 24 nonconserved intragenic SB‐miRNA genes, and 23 nonconserved intragenic Va‐miRNA genes were labeled on the basis of their locations in host genes, and 15 MIRNA clusters were detected. Interestingly, five miRNA pairs, namely, Vv‐MIR395b and Vv‐MIR395c, Vv‐MIR482 and Vv‐MIRC13, Vv‐MIR172a and Va‐MIR057, SB‐MIR024 and Vv‐MIRC35, and Vv‐MIRC36 and Va‐MIR073 were clustered in the host genes GSVIVT01011558001, GSVIVT01008132001, GSVIVT01031524001, GSVIVT01028156001, and GSVIVT01024516001, respectively. To validate the existence of target genes and miRNA‐guided cleavage sites, 3′‐end product of four predicted target messenger RNAs were amplified by RNA ligase‐mediated 5′ rapid amplification of cDNA ends. In addition, we also conducted contrastive analysis on the genomic location of miRNAs and their potential target genes. Results showed that the order of priority of miRNA–target interaction may be less closely related with their genomic location. These findings could benefit some further study on grapevine functional genomics and will provide new insights into the regulatory mechanisms and evolution of miRNAs in Vitis species.

Grapevine immune signaling network in response to drought stress as revealed by transcriptomic analysis

Drought is a ubiquitous abiotic factor that severely impedes growth and development of horticulture crops. The challenge postured by global climate change is the evolution of drought-tolerant cultivars that could cope with concurrent stress. Hence, in this study, biochemical, physiological and transcriptome analysis were investigated in drought-treated grapevine leaves. The results revealed that photosynthetic activity and reducing sugars were significantly diminished which were positively correlated with low stomatal conductance and CO2 exchange in drought-stressed leaves. Further, the activities of superoxide dismutase, peroxidase, and catalase were significantly actuated in the drought-responsive grapevine leaves. Similarly, the levels of abscisic acid and jasmonic acid were also significantly increased in the drought-stressed leaves. In transcriptome analysis, 12,451 differentially-expressed genes (DEGs) were annotated, out of which 8021 DEGs were up-regulated and 4430 DEGs were down-regulated in response to drought stress. In addition, the genes encoding pathogen-associated molecular pattern (PAMP) triggered immunity (PTI), including calcium signals, protein phosphatase 2C, calcineurin B-like proteins, MAPKs, and phosphorylation (FLS2 and MEKK1) cascades were up-regulated in response to drought stress. Several genes related to plant-pathogen interaction pathway (RPM1, PBS1, RPS5, RIN4, MIN7, PR1, and WRKYs) were also found up-regulated in response to drought stress. Overall the results of present study showed the dynamic interaction of DEG in grapevine physiology which provides the premise for selection of defense-related genes against drought stress for subsequent grapevine breeding programs.

Role of microRNAs and their target genes in salinity response in plants

Salinity is one of the most common environmental stresses that affects plant growth and development. Plants employ complex mechanisms of gene regulation to respond salinity stress. MicroRNAs (miRNAs) are small noncoding RNAs with important regulatory roles in plant growth and development. Furthermore, they are also considered as critical cell elements orchestrating plants response to stress conditions. Many approaches, such as molecular cloning and high-throughput sequencing using computational tools have been adapted to profile miRNA expression patterns during abiotic stresses. In fact, the identification of these master regulators could pave the way towards better understanding of the molecular regulation of stress adaptation in plants. Many target genes of miRNAs are transcription factors which further regulate a set of downstream genes to have effects on physiological responses. This review summarizes recent molecular studies on the regulatory roles of plant miRNAs and their target genes in plant metabolism under salt stress conditions.

Characterization of VvPAL-like promoter from grapevine using transgenic tobacco plants.

A 2000-bp 5′-flanking region of VvPAL-like was isolated from ‘Summer Black’ grapevine by PCR amplification, named pVvPAL-like. To gain a better understanding of the expression and regulatory mechanism of VvPAL-like, a chimeric expression unit consisting of the β-glucuronidase (GUS) reporter gene under the control of a 2000-bp fragment of the VvPAL-like promoter was transformed into tobacco via Agrobacterium tumefaciens. Histochemical staining showed that the full-length promoter directs efficient expression of the reporter gene in cotyledons and hypocotyls, stigma, style, anthers, pollen, ovary, trichomes, and vascular bundles of transgenic plants. A series of 5′ progressive deletions of the promoter revealed the presence of a negative regulatory region (−424 to −292) in the VvPAL-like promoter. Exposure of the transgenic tobacco plants to various abiotic stresses demonstrated that the full-length construct could be induced by light, copper (Cu), abscisic acid (ABA), indole-3-acetic (IAA), methyl jasmonate (MeJA) (N-1-naphthylphthalamic acid), ethylene, and drought. Furthermore, the ethylene-responsive region was found to be located in the −1461/−930 fragment, while the element(s) for the MeJA-responsive expression may be present in the −424/−292 region in the VvPAL-like promoter. These findings will help us to better understand the molecular mechanisms by which VvPAL-like participates in biosynthesis of flavonoids and stress responses.

Analysis of the regulation networks in grapevine reveals response to waterlogging stress and candidate gene-marker selection for damage severity.

Owing to the climate change impacts, waterlogging is one of the most hazardous abiotic stresses to crops, which also can result in a serious reduction in the quantity and quality of grape berry and wine production during the rainy season. Therefore, the exploration of the response mechanism of grape to waterlogging is necessary, for which the analysis of the transcriptomic regulation networks of grapevine leaves in response to waterlogging stress was carried out. In this study, 12 634 genes were detected in both waterlogging stress and control grapevine plants, out of which 6837 genes were differentially expressed. A comparative analysis revealed that genes functioning in the antioxidant system, glycolysis and fermentation pathway, chlorophyll metabolism, amino acid metabolism and hormones were activated to reduce injury to grapes under the waterlogging stress. Meanwhile, genes encoding class-2 non-symbiotic haemoglobin were determined as important in waterlogging acclimation. Additionally, the expression variations of three marker genes were found to be informative and can be used to predict the viability of the grapevines subjected to waterlogging. This research not only probes the molecular mechanism underlying grapevine waterlogging tolerance but also puts forward an idea about the application of gene expression information to practical management.

In silico identification and computational characterization of endogenous small interfering RNAs from diverse grapevine tissues and stages

Small interfering RNAs (siRNAs) are effectors of regulatory pathways underlying plant development, metabolism, and stress- and nutrient-signaling regulatory networks. The endogenous siRNAs are generally not conserved between plants; consequently, it is necessary and important to identify and characterize siRNAs from various plants. To address the nature and functions of siRNAs, and understand the biological roles of the huge siRNA population in grapevine (Vitis vinifera L.). The high-throughput sequencing technology was used to identify a large set of putative endogenous siRNAs from six grapevine tissues/organs. Subsequently, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis was performed to classify the target genes of siRNA. In total, 520,519 candidate siRNAs were identified and their expression profiles exhibited typical temporal characters during grapevine development. In addition, we identified two grapevine trans-acting siRNA (TAS) gene homologs (VvTAS3 and VvTAS4) and the derived trans-acting siRNAs (tasiRNAs) that could target grapevine auxin response factor (ARF) and myeloblastosis (MYB) genes. Furthermore, the GO and KEGG analysis of target genes showed that most of them covered a broad range of functional categories, especially involving in disease-resistance process. The large-scale and completely genome-wide level identification and characterization of grapevine endogenous siRNAs from the diverse tissues by high throughput technology revealed the nature and functions of siRNAs in grapevine.

Genome-Wide Identification of PIFs in Grapes (Vitis vinifera L.) and Their Transcriptional Analysis under Lighting/Shading Conditions

Phytochrome-interacting factors (PIFs), as the basic helix–loop–helix (bHLH) transcription factors, are the primary signaling partners for phytochromes (PHY) that play a key role in PHY-mediated light signal transduction. At present, there are few studies on PIFs in fruit trees. In order to clarify the status of PIFs in grapevines, we identified members of the grape PIFs family and conducted phylogenetic and expression analysis. We identified PIF1, PIF3, PIF4, and PIF7 in PIFs families of the grapevine (Vitis vinifera L.), which were distributed on four different chromosomes with similar gene structures. Except for the closer relationship with PIF1 of citrus, PIFs of grape were distant from the other fruit species such as apple, pear, peach, and strawberry. The VvPIFs (except VvPIF4) were located in the syntenic block with those from Arabidopsis thaliana, Solanum lycopersicum, or Citrus sinensis. In addition to PIF1, all PIFs in grapevines have conserved active PHYB binding (APB) sequences. VvPIF1 has a conserved PIF1-specific active PHYA binding (APA) sequence, while amino acid mutations occurred in the specific APA sequence in VvPIF3. Interestingly, two specific motifs were found in the PIF4 amino acid sequence. The photoreceptor-related elements in the VvPIFs promoter region were the most abundant. PIF1, LONG HYPOCOTYL 5 (HY5) and PIF3, PIF4, GIBBERELLIC ACID INSENSITIVE 1 (GAI1) may interact with each other and participate together in light signal transduction. The relative expression levels of the VvPIFs showed diverse patterns in the various organs at different developmental stages, of which PIF4 was most highly expressed. Prior to maturation, the expression of PIF4 and PIF7 in the skin of the different cultivars increased, while the expression of all PIFs in the flesh decreased. The transcription level of PIFs in grape leaves was sensitive to changes in lighting and shading. Shading treatment was beneficial for enhancing the transcription level of VvPIFs, but the effect on VvPIF3 and VvPIF4 was time-controlled. We concluded that PIFs in grapevines are both conservative and species-specific. The identification and analysis of grape PIFs could provide a theoretical foundation for the further construction of grape light regulation networks.