Drought is a major environmental factor that significantly limits crop yield and quality worldwide. Basic helix-loop-helix (bHLH) transcription factors have been reported to participate in the ...regulation of various abiotic stresses. In this study, a bHLH transcription factor in apple, MdbHLH130, which contains a highly conserved bHLH domain, was isolated and characterized. qRT-PCR and PMdbHLH130::GUS analyses showed that MdbHLH130 was notably induced in response to dehydration stress. Compared with the wild-type (WT), transgenic apple calli overexpressing MdbHLH130 displayed greater resistance to PEG6000 treatment. In contrast, the MdbHLH130-Anti lines were more sensitive to PEG6000 treatment than WT. Moreover, ectopic expression of MdbHLH130 in tobacco improved tolerance to water deficit stress, and plants exhibited higher germination rates and survival rates, longer roots, and lower ABA-induced stomatal closure and leaf water loss than the WT control. Furthermore, overexpression of MdbHLH130 in tobacco also led to lower electrolyte leakage, malondialdehyde contents, and reactive oxygen species (ROS) accumulation and upregulation of the expression of some ROS-scavenging and stress-responsive genes under water deficit stress. In addition, MdbHLH130 transgenic tobacco plants exhibited improved tolerance to oxidative stress compared with WT. In conclusion, these results indicate that MdbHLH130 acts as a positive regulator of water stress responses through modulating stomatal closure and ROS-scavenging in tobacco.
ABSTRACTGovernment service hotlines are closely related to people’s lives, and the hotline events assignment task is very important in China, which affects people’s satisfaction with the Chinese ...government. To address the challenge of improving hotline event assignment accuracy, which is currently hindered by the lack of prior knowledge in the traditional direct departmental assignment approach, we introduce a novel hotline event assignment model based on Knowledge Base Question Answering (KBQA) to leverage prior knowledge and enhance assignment performance. The model extracts event key information by using event extraction module, and the key information of event extracte by this module is used to construct historical events knowledge graph. Then we employ subgraph retrieval module and text retrieval module to obtain relevant prior knowledge from the knowledge graph and “sanding” text respectively. The integrated prior knowledge is then used to predict department scores, guiding the selection of the optimal assignment department through ranking. The experiment results show that the proposed method outperforms existing baseline methods. Meanwhile, the Knowledge Graph (KG) edge elimination experiment also shows that the proposed model is better than other baseline models in terms of incomplete KG.
The plant-specific TCP transcription factors play different functions in multiple processes of plant growth and development. TCP family genes have been identified in several plant species, but no ...comprehensive analysis of the TCP family in grapevine has been undertaken to date, especially their roles in fruit development.
A total of 18 non-redundant grapevine TCP (VvTCP) genes distributing on 11 chromosomes were identified. Phylogenetic and structural analysis showed that VvTCP genes were divided into two main classes - class I and class II. The Class II genes were further classified into two subclasses, the CIN subclass and the CYC/TB1 subclass. Segmental duplication was a predominant duplication event which caused the expansion of VvTCP genes. The cis-acting elements analysis and tissue-specific expression patterns of VvTCP genes demonstrated that these VvTCP genes might play important roles in plant growth and development. Expression patterns of VvTCP genes during fruit development and ripening were analyzed by RNA-Seq and qRT-PCR. Among them, 11 VvTCP genes were down-regulated during different fruit developmental stages, while only one VvTCP genes were up-regulated, suggesting that most VvTCP genes were probably related to early development in grapevine fruit. Futhermore, the expression of most VvTCP genes can be inhibited by drought and waterlogging stresses.
Our study establishes the first genome-wide analysis of the grapevine TCP gene family and provides valuable information for understanding the classification and functions of the TCP genes in grapevine.
The B-BOX (BBX) proteins are the class of zinc-finger transcription factors and can regulate plant growth, development, and endure stress response. In plants, the BBX gene family has been identified ...in Arabidopsis, rice, and tomato. However, no systematic analysis of BBX genes has been undertaken in grapevine.
In this study, 24 grapevine BBX (VvBBX) genes were identified by comprehensive bioinformatics analysis. Subsequently, the chromosomal localizations, gene structure, conserved domains, phylogenetic relationship, gene duplication, and cis-acting elements were analyzed. Phylogenetic analysis divided VvBBX genes into five subgroups. Numerous cis-acting elements related to plant development, hormone and/or stress responses were identified in the promoter of the VvBBX genes. The tissue-specific expressional dynamics of VvBBX genes demonstrated that VvBBXs might play important role in plant growth and development. The transcript analysis from transcriptome data and qRT-PCR inferred that 11 VvBBX genes were down-regulated in different fruit developmental stages, while three VvBBX genes were up-regulated. It is also speculated that VvBBX genes might be involved in multiple hormone signaling (ABA, ethylene, GA3, and CPPU) as transcriptional regulators to modulate berry development and ripening. VvBBX22 seems to be responsive to multiple hormone signaling, including ABA, ethylene GA3, and CPPU. Some VvBBX genes were strongly induced by Cu, salt, waterlogging, and drought stress treatment. Furthermore, the expression of VvBBX22 proposed its involvement in multiple functions, including leaf senescence, abiotic stress responses, fruit development, and hormone response.
Our results will provide the reference for functional studies of BBX gene family, and highlight its functions in grapevine berry development and ripening. The results will help us to better understand the complexity of the BBX gene family in abiotic stress tolerance and provide valuable information for future functional characterization of specific genes in grapevine.
Apple bitter pit primarily occurs during fruit ripening and storage; however, its formation mechanism remains unclear. Although it is considered that Ca
2+
deficiency causes metabolic disorders in ...apples, there have been few studies on the mechanism of the bitter pit from the perspective of cell structure. At the fruit ripening stage, the fruit with a bitter pit on the tree was taken as the research material. In this study, the microscopic observation revealed numerous amyloplasts in the pulp cells of apples affected with bitter pit, but not in the healthy pulp. Furthermore, the results of fluorescence staining and transmission electron microscopy (TEM) revealed that the bitter pit pulp cells undergo programmed cell death (PCD), their nuclear chromosomes condense, and amyloplast forms autophagy. The cytoplasmic Ca
2+
concentration in the healthy fruits was lowest near the peduncle, followed by that in the calyx, whereas it was highest at the equator. In contrast, the cytoplasmic Ca
2+
concentration in apple fruits showing bitter pit disorder was lowest near the peduncle and highest in the calyx. Moreover, the cytosolic Ca
2+
concentration in the flesh cells of apples with the bitter pit was much lower than that in the healthy apple flesh cells; however, the concentration of Ca
2+
in the vacuoles of fruits with the bitter pit was higher than that in the vacuoles of healthy fruits. In summary, bitter pit pulp cells contain a large number of amyloplasts, which disrupts the distribution of Ca
2+
in the pulp cells and causes PCD. These two processes lead to an imbalance in cell metabolism and induce the formation of a bitter pit.
The most direct technique for studying calcium, which is an essential element for pollen tube growth, is Ca2+ imaging. Because membranes are relatively impermeable, the loading of fluorescent Ca2+ ...probes into plant cells is a challenging task. Thus, we have developed a new method of loading fluo-4 acetoxymethyl ester into cells that uses a cell lysis solution to improve the introduction of this fluorescent dye into pollen tubes. Using this method, the loading times were reduced to 15 min. Furthermore, loading did not have to be performed at low (4°C) temperatures and was successful at room temperature, and pluronic F-127 was not required, which would theoretically allow for the loading of an unlimited number of cells. Moreover, the method can also be used to fluorescently stain root hairs.
Adventitious root formation is essential for plant propagation, development, and response to various stresses. Reactive oxygen species (ROS) are essential for adventitious root formation. However, ...information on Respiratory Burst Oxidase Homolog (RBOH), a key enzyme that catalyzes the production ROS, remains limited in woody plants. Here, a total of 44 RBOH genes were identified from six Rosaceae species (Malus domestica, Prunus avium, Prunus dulcis 'Texas', Rubus occidentalis, Fragaria vesca and Rosa chinensis), including ten from M. domestica. Their phylogenetic relationships, conserved motifs and gene structures were analyzed. Exogenous treatment with the RBOH protein inhibitor diphenyleneiodonium (DPI) completely inhibited adventitious root formation, whereas exogenous H.sub.2 O.sub.2 treatment enhanced adventitious root formation. In addition, we found that ROS accumulated during adventitious root primordium inducing process. The expression levels of MdRBOH-H, MdRBOH-J, MdRBOH-A, MdRBOH-E1 and MdRBOH-K increased more than two-fold at days 3 or 9 after auxin treatment. In addition, cis-acting element analysis revealed that the MdRBOH-E1 promoter contained an auxin-responsive element and the MdRBOH-K promoter contained a meristem expression element. Based on the combined results from exogenous DPI and H.sub.2 O.sub.2 treatment, spatiotemporal expression profiling, and cis-element analysis, MdRBOH-E1 and MdRBOH-K appear to be candidates for the control of adventitious rooting in apple.
Among the Rosaceae species, the gametophytic self-incompatibility (GSI) is controlled by a single multi-allelic S locus, which is composed of the pistil-S and pollen-S genes. The pistil-S gene ...encodes a polymorphic ribonuclease (S-RNase), which is essential for identifying self-pollen. However, the S-RNase system has not been fully characterized. In this study, the self-S-RNase inhibited the Ca
-permeable channel activity at pollen tube apices and the selectively decreased phospholipase C (
) activity in the plasma membrane of
pollen tubes. Self-S-RNase decreased the Ca
influx through a
-mediated signaling pathway. Phosphatidylinositol-specific
has a 26-amino acid insertion in pollen tubes of the 'Jinzhuili' cultivar, which is a spontaneous self-compatible mutant of the 'Yali' cultivar. 'Yali' plants exhibit a typical S-RNase-based GSI. Upon self-pollination,
gene expression is significantly higher in 'Jinzhuili' pollen tubes than that in 'Yali' pollen tubes. Moreover, the
in pollen tubes can only interact with one of the two types of S-RNase from the style. In the
Rehd, the
directly interacted with the S
-RNase in the pollen tube, but not with the S
-RNase. Collectively, our results reveal that the effects of S-RNase on
activity are required for S-specific pollen rejection, and that
-IP
participates in the self-incompatibility reaction of
species.
The 14-3-3 (GRF, general regulatory factor) regulatory proteins are highly conserved and are widely distributed throughout the eukaryotes. They are involved in the growth and development of organisms
...target protein interactions. Although many plant 14-3-3 proteins were identified in response to stresses, little is known about their involvement in salt tolerance in apples. In our study, nineteen apple 14-3-3 proteins were cloned and identified. The transcript levels of
genes were either up or down-regulated in response to salinity treatments. Specifically, the transcript level of
(a member of the
genes family) decreased due to salt stress treatment. The phenotypes of transgenic tobacco lines and wild-type (WT) did not affect plant growth under normal conditions. However, the germination rate and salt tolerance of transgenic tobacco was lower compared to the WT. Transgenic tobacco demonstrated decreased salt tolerance. The transgenic apple calli overexpressing
exhibited greater sensitivity to salt stress compared to the WT plants, whereas the
transgenic apple calli improved salt stress tolerance. Moreover, the salt stress-related genes (
,
,
,
,
,
,
, and
) were more strongly down-regulated in
transgenic apple calli lines than in the WT when subjected to salt stress treatment. Taken together, these results provide new insights into the roles of 14-3-3 protein MdGRF6 in modulating salt responses in plants.
Abstract
Cytosolic Ca
2+
plays a key role in signal transduction in plants. Calcium imaging is the most common approach to studying dynamic changes in the cytoplasmic Ca
2+
content. Here, we used ...mature ‘Fuji’ apples (
Malus pumila Mill
.) to obtain viable protoplasts from flesh tissue cells by enzymatic hydrolysis; then, three small-molecule fluorescent probes (fluo-8/AM, fluo-4/AM, and rhod-2/AM) were loaded into the protoplasts. All three Ca
2+
fluorescent probes successfully entered the cytoplasm but did not enter the vacuole. Both the Ca
2+
chelator (EGTA) and Ca
2+
channel blocker (La
3+
) reduced the fluorescence intensity in the cytoplasm. The calcium ionophore A23187 increased the fluorescence intensity in the cytoplasm at 5 µmol/L but decreased it at 50 µmol/L. Additionally, A23187 reversed the fluorescence intensity in the cytoplasm, which was decreased by La
3+
. Ionomycin is also a calcium ionophore that can increase the fluorescence intensity of calcium in the cytoplasm. These results suggest that small-molecule Ca
2+
fluorescent probes can be used to detect changes in cytosolic calcium levels in the cells of fruit flesh tissue. In addition, the optimum concentration of fluo-8/AM was determined to be 5 µmol/L. This was the first time that protoplasts have been isolated from apple flesh tissue cells and small-molecule fluorescent probes have been used to detect calcium in the cytoplasm of flesh tissue cells. This study provides a new method to study calcium signal transduction in fruit flesh tissue.
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