ABSTRACT
The characterization of the PYL/RCAR ABA receptors in a great deal of plant species has dramatically advanced the study of ABA functions involved in key physiological processes. However, the genes in this family are still unclear in Lycium (Goji) plants, one of the well-known economically, medicinally, and ecologically valuable fruit crops. In the present work, 12 homologs of Arabidopsis PYL/RCAR ABA receptors were first identified and characterized from Lycium (L.) barbarum (LbPYLs). The quantitative real-time PCR (qRT-PCR) analysis showed that these genes had clear tissue-specific expression patterns, and most of them were transcribed in the root with the largest amount. Among the three subfamilies, while the Group I and Group III members were down-regulated by extraneous ABA, the Group II members were up-regulated. At 42 °C, most transcripts showed a rapid and violent up-regulation response to higher temperature, especially members of Group II. One of the genes in the Group II members, LbPYL10, was further functionally validated by virus-induced gene silencing (VIGS) technology. LbPYL10 positively regulates heat stress tolerance in L. barbarum by alleviating chlorophyll degradation, thus maintaining chlorophyll stability. Integrating the endogenous ABA level increase following heat stress, it may be concluded that LbPYL-mediated ABA signaling plays a vital role in the thermotolerance of L. barbarum plants. Our results highlight the strong potential of LbPYL genes in breeding genetically modified L. barbarum crops that acclimate to climate change.
ABSTRACT
Plant−water relations mediated by aquaporins (AQPs) play vital roles in both key plant growth processes and responses to environmental challenges. As a well-known medicinal and edible plant, the harsh natural growth habitat endows Lycium plants with ideal materials for stress biology research. However, the details of their molecular switch for water transport remain unclear. In the present work, we first identified and characterized AQP family genes from Lycium (L.) barbarum at the genome scale and conducted systemic bioinformatics and expression analyses. The results showed that there were 38 Lycium barbarum AQPs (LbAQPs) in L. barbarum, which were classified into four subfamilies, including 17 LbPIP, 9 LbTIP, 10 LbNIP, and 2 LbXIP. Their encoded genes were unevenly distributed on all 12 chromosomes, except chromosome 10. Three of these genes encoded truncated proteins and three genes underwent clear gene duplication events. Cis-acting element analysis indicated that the expression of LbAQPs may be mainly regulated by biotic/abiotic stress, phytohormones and light. The qRT-PCR assay indicated that this family of genes presented a clear tissue-specific expression pattern, in which most of the genes had maximal transcript levels in roots, stems, and leaves, while there were relatively lower levels in flowers and fruits. Most of the LbAQP genes were downregulated during L. barbarum fruit ripening and presented a negative correlation with the fruit relative water content (RWC). Most of their transcripts presented a quick and sharp upregulation response to heat stress following exposure of the 2-month-old seedlings to a 42 °C temperature for 0, 1, 3, 12, or 24 h. Our results proposed that LbAQPs were involved in L. barbarum key development events and abiotic stress responses, which may lay a foundation for further studying the molecular mechanism of the water relationship of Lycium plants, especially in harsh environments.
ABSTRACT
Anthocyanin-derived fleshy fruit pigmentation has become an excellent system for studying the regulatory network underlying fruit ripening and quality. The transcriptional control of anthocyanin biosynthesis by MYB-bHLH-WDR complexes has been well established, but the intermediate signals through which the environmental or developmental cues regulate these transcription factors remain poorly understood. Here we found that nitric oxide (NO) production during Lycium fruit ripening decreased progressively presenting a negative relationship with anthocyanins. After cloning of the nitric reductase (NR) gene from Lycium barbarum (LbNR) plants, we demonstrated that LbNR-derived NO partially inhibited anthocyanin biosynthesis but enhanced proanthocyanidin (PA) accumulation, and delayed fruit coloration. Application of the NO donor, sodium nitroprusside (SNP), produced a similar effect. The endogenous or exogenous NO downregulated the transcripts both of the regulatory genes and the structural genes that related to anthocyanin biosynthesis, while upregulated both of those genes that related to PA biosynthesis. Given there is a significant negative relationship between the levels of anthocyanins and PAs during Lycium fruit ripening, NO not only inhibited anthocyanin de novo biosynthesis but redirected the flavonoid biosynthetic pathway from anthocyanins to PA production. Two types of LrMYB transcription factors of opposite nature, namely anthocyanin-specific and PA-specific, which belong to the R2R3-MYB subfamily and 1R-MYB subfamily, respectively, were identified from L. ruthenicum fruits. It was further found that NO acts by antagonizing the ABA signaling, a phytohormone we have previously shown playing a positive role in Lycium fruit coloration. Our results provided particularly novel information about NO-ABA-anthocyanin interplay during Lycium fruit development and ripening, which may fill a gap between the developmental cues and the transcriptional regulation of anthocyanin biosynthesis.
ABSTRACT
BACKGROUND: Anthocyanins, which are colored pigments, have long been used as food and pharmaceutical ingredients due to their potential health benefits, but the intermediate signals through which environmental or developmental cues regulate anthocyanin biosynthesis remains poorly understood. Fleshy fruits have become a good system for studying the regulation of anthocyanin biosynthesis, and exploring the mechanism underlying pigment metabolism is valuable for controlling fruit ripening. RESULTS: The present study revealed that ABA accumulated during Lycium fruit ripening, and this accumulation was positively correlated with the anthocyanin contents and the LbNCED1 transcript levels. The application of exogenous ABA and of the ABA biosynthesis inhibitor fluridon increased and decreased the content of anthocyanins in Lycium fruit, respectively. This is the first report to show that ABA promotes the accumulation of anthocyanins in Lycium fruits. The variations in the anthocyanin content were consistent with the variations in the expression of the genes encoding the MYB-bHLH-WD40 transcription factor complex or anthocyanin biosynthesis-related enzymes. Virus-induced LbNCED1 gene silencing significantly slowed fruit coloration and decreased both anthocyanin and ABA accumulation during Lycium fruit ripening. An qRT-PCR analysis showed that LbNCED1 gene silencing clearly reduced the transcript levels of both structural and regulatory genes in the flavonoid biosynthetic pathway. CONCLUSIONS: Based on the results, a model of ABA-mediated development-dependent anthocyanin biosynthesis and fruit coloration during Lycium fruit maturation was proposed. In this model, the developmental cues transcriptionally activates LbNCED1 and thus enhances accumulation of the phytohormone ABA, and the accumulated ABA stimulates transcription of the MYB-bHLH-WD40 transcription factor complex to upregulate the expression of structural genes in the flavonoid biosynthetic pathway and thereby promoting anthocyanin production and fruit coloration. Our results provide a valuable strategy that could be used in practice to regulate the ripening and quality of fresh fruit in medicinal and edible plants by modifying the phytohormone ABA.
Subject(s)
Abscisic Acid/metabolism , Anthocyanins/biosynthesis , Fruit/metabolism , Lycium/metabolism , Pigmentation , Plant Growth Regulators/metabolism , Dioxygenases/genetics , Fruit/growth & development , Gene Expression Regulation, Plant , Gene Silencing , Genes, Plant , Lycium/genetics , Lycium/growth & development , Plant Proteins/genetics , Signal TransductionABSTRACT
PYR/PYLs function as ABA receptors and are key regulators during plant drought stress response. Previously we screened drought tolerance of Arabidopsis ABA receptors PYR/PYLs under the control of five different promoters. In this study, we characterized drought stress tolerance of AtPYL5 transgene under the control of one guard cell specific promoter, pGC1. pGC1::AtPYL5 transgenic Arabidopsis exhibited reduced transpiration rate and decreased water loss after drought treatment. Transformation of pGC1::AtPYL5 in Arabidopsis also decreased oxidative stress damage and improved photosynthesis under drought stress condition. These results indicated that pGC1::AtPYL5 construct is effective and might pave new way to develop genetically engineered plants to improve drought stress tolerance.
Subject(s)
Arabidopsis Proteins/physiology , Arabidopsis/metabolism , Intracellular Signaling Peptides and Proteins/physiology , Antioxidants/metabolism , Arabidopsis/physiology , Arabidopsis Proteins/metabolism , Dehydration , Hydrogen Peroxide/metabolism , Intracellular Signaling Peptides and Proteins/metabolism , Oxidative Stress , Photosynthesis , Plant Leaves/metabolism , Plant Transpiration , Plants, Genetically Modified , Promoter Regions, Genetic , Water/metabolismABSTRACT
Following publication of the original article [1], a reader spotted that the article appears to have some misplaced/duplicated figures. In particular, Fig. 5a and Fig. 6a appear to be identical, and do not match what is written in the text. The authors apologized for this oversight and supplied the original pictures, which are reproduced below.
ABSTRACT
From the different functions ABA exerted between the aboveground and belowground, seed and vegetative tissues, primary root and lateral root, stimulating stomatal closure and inhibiting stomatal opening, between young and senescence leaves in stomatal movement, among different cells in plasma membrane water permeability, we addressed the organ-, tissue-, cell-, physiological processes-, and development stage specificities of PYR1/PYL/RCAR ABA receptors. This specificity may reflect the spatio-temporal properties of water potentials as well as the endogenous ABA levels in detail context, which plus the various affinities among this receptor families, resulted in the specificity of the transcripts as well as genes functions. PYR1/PYL/RCAR ABA receptors may integrate the message of ABA resource (local signaling or long distance signaling) and concentration, thus fine-tuning ABA response to environmental- and developmental cues. It also evolutionally affording land plants sophisticated mechanism to survival adverse environments.
Subject(s)
Abscisic Acid/metabolism , Arabidopsis Proteins/metabolism , Arabidopsis/metabolism , Carrier Proteins/metabolism , Arabidopsis/genetics , Arabidopsis/physiology , Arabidopsis Proteins/genetics , Carrier Proteins/genetics , Membrane Transport Proteins/genetics , Membrane Transport Proteins/metabolismABSTRACT
BACKGROUND: The different actions of abscisic acid (ABA) in the aboveground and belowground parts of plants suggest the existence of a distinct perception mechanism between these organs. Although characterization of the soluble ABA receptors PYR1/PYL/RCAR as well as core signaling components has greatly advanced our understanding of ABA perception, signal transduction, and responses, the environment-dependent organ-specific sensitivity of plants to ABA is less well understood. RESULTS: By performing real-time quantitative PCR assays, we comprehensively compared transcriptional differences of core ABA signaling components in response to ABA or osmotic/dehydration stress between maize (Zea mays L.) roots and leaves. Our results demonstrated up-regulation of the transcript levels of ZmPYLs homologous to dimeric-type Arabidopsis ABA receptors by ABA in maize primary roots, whereas those of ZmPYLs homologous to monomeric-type Arabidopsis ABA receptors were down-regulated. However, this trend was reversed in the leaves of plants treated with ABA via the root medium. Although the mRNA levels of ZmPYL1-3 increased significantly in roots subjected to polyethylene glycol (PEG)-induced osmotic stress, ZmPYL4-11 transcripts were either maintained at a stable level or increased only slightly. In detached leaves subjected to dehydration, the transcripts of ZmPYL1-3 together with ZmPYL5, ZmPYL6, ZmPYL10 and ZmPYL11 were decreased, whereas those of ZmPYL4, ZmPYL7 and ZmPYL8 were significantly increased. Our results also showed that all of the evaluated transcripts of PP2Cs and SnRK2 were quickly up-regulated in roots by ABA or osmotic stress; conversely they were either up-regulated or maintained at a constant level in leaves, depending on the isoforms within each family. CONCLUSIONS: There is a distinct profile of PYR/PYL/RCAR ABA receptor gene expression between maize roots and leaves, suggesting that monomeric-type ABA receptors are mainly involved in the transmission of ABA signals in roots but that dimeric-type ABA receptors primarily carry out this function in leaves. Given that ZmPYL1 and ZmPYL4 exhibit similar transcript abundance under normal conditions, our findings may represent a novel mechanism for species-specific regulation of PYR/PYL/RCAR ABA receptor gene expression. A difference in the preference for core signaling components in the presence of exogenous ABA versus stress-induced endogenous ABA was observed in both leaves and roots. It appears that core ABA signaling components perform their osmotic/dehydration stress response functions in a stress intensity-, duration-, species-, organ-, and isoform-specific manner, leading to plasticity in response to adverse conditions and, thus, acclimation to life on land. These results deepen our understanding of the diverse biological effects of ABA between plant leaves and roots in response to abiotic stress at the stimulus-perception level.
Subject(s)
Abscisic Acid/pharmacology , Gene Expression Regulation, Plant/drug effects , Plant Leaves/genetics , Plant Proteins/genetics , Plant Roots/genetics , Receptors, Cell Surface/genetics , Seedlings/genetics , Zea mays/genetics , Abscisic Acid/metabolism , Arabidopsis/genetics , Dehydration , Dose-Response Relationship, Drug , Gene Expression Profiling/methods , Phylogeny , Plant Growth Regulators/metabolism , Plant Growth Regulators/pharmacology , Plant Proteins/classification , Polyethylene Glycols/pharmacology , Receptors, Cell Surface/classification , Reverse Transcriptase Polymerase Chain Reaction , Time Factors , Up-Regulation/drug effectsABSTRACT
Drought stress is an important environmental factor limiting plant productivity. In this study, we screened drought-resistant transgenic plants from 65 promoter-pyrabactin resistance 1-like (PYL) abscisic acid (ABA) receptor gene combinations and discovered that pRD29A::PYL9 transgenic lines showed dramatically increased drought resistance and drought-induced leaf senescence in both Arabidopsis and rice. Previous studies suggested that ABA promotes senescence by causing ethylene production. However, we found that ABA promotes leaf senescence in an ethylene-independent manner by activating sucrose nonfermenting 1-related protein kinase 2s (SnRK2s), which subsequently phosphorylate ABA-responsive element-binding factors (ABFs) and Related to ABA-Insensitive 3/VP1 (RAV1) transcription factors. The phosphorylated ABFs and RAV1 up-regulate the expression of senescence-associated genes, partly by up-regulating the expression of Oresara 1. The pyl9 and ABA-insensitive 1-1 single mutants, pyl8-1pyl9 double mutant, and snrk2.2/3/6 triple mutant showed reduced ABA-induced leaf senescence relative to the WT, whereas pRD29A::PYL9 transgenic plants showed enhanced ABA-induced leaf senescence. We found that leaf senescence may benefit drought resistance by helping to generate an osmotic potential gradient, which is increased in pRD29A::PYL9 transgenic plants and causes water to preferentially flow to developing tissues. Our results uncover the molecular mechanism of ABA-induced leaf senescence and suggest an important role of PYL9 and leaf senescence in promoting resistance to extreme drought stress.
Subject(s)
Abscisic Acid/metabolism , Arabidopsis Proteins/physiology , Arabidopsis/physiology , Carrier Proteins/physiology , Droughts , Plant Leaves/physiology , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Carrier Proteins/genetics , Carrier Proteins/metabolism , Intracellular Signaling Peptides and Proteins , Oryza/genetics , Oryza/physiology , Phosphorylation , Plants, Genetically Modified , Signal Transduction , Stress, PhysiologicalABSTRACT
The C-REPEAT-BINDING FACTOR (CBF) pathway has important roles in plant responses to cold stress. How the CBF genes themselves are activated after cold acclimation remains poorly understood. In this study, we characterized cold tolerance of null mutant of RNA-DIRECTED DNA METHYLATION 4 (RDM4), which encodes a protein that associates with RNA polymerases Pol V and Pol II, and is required for RNA-directed DNA methylation (RdDM) in Arabidopsis. The results showed that dysfunction of RDM4 reduced cold tolerance, as evidenced by decreased survival and increased electrolyte leakage. Mutation of RDM4 resulted in extensive transcriptomic reprogramming. CBFs and CBF regulon genes were down-regulated in rdm4 but not nrpe1 (the largest subunit of PolV) mutants, suggesting that the role of RDM4 in cold stress responses is independent of the RdDM pathway. Overexpression of RDM4 constitutively increased the expression of CBFs and regulon genes and decreased cold-induced membrane injury. A great proportion of genes affected by rdm4 overlapped with those affected by CBFs. Chromatin immunoprecipitation results suggested that RDM4 is important for Pol II occupancy at the promoters of CBF2 and CBF3. We present evidence of a considerable role for RDM4 in regulating gene expression at low temperature, including the CBF pathway in Arabidopsis.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/physiology , Cold Temperature , Stress, Physiological , Trans-Activators/metabolism , Transcription Factors/metabolism , Arabidopsis/genetics , Arabidopsis Proteins/genetics , Cell Membrane/metabolism , Electrolytes/metabolism , Freezing , Gene Expression Profiling , Gene Expression Regulation, Plant , Gene Ontology , Genes, Plant , Mutation/genetics , Oxidative Stress/genetics , Plant Leaves/physiology , Promoter Regions, Genetic/genetics , Protein Binding , RNA Polymerase II/metabolism , Reactive Oxygen Species/metabolism , Seedlings/metabolism , Stress, Physiological/genetics , Trans-Activators/genetics , Transcription Factors/geneticsABSTRACT
Pyrabactin, an agonist of abscisic acid (ABA), has led to the isolation and characterization of pyrabactin resistance 1/pyrabactin resistance 1-like (PYR1/PYLs) ABA receptors in Arabidopsis, which has well explained ABA-mediated stomatal movement and stress-related gene expression. In addition to inducing stomatal closure and inhibiting transpiration, ABA can also enhance root hydraulic conductivity (Lpr), thus maintaining water balance under water deficiency-related stress, but its molecular mechanism remains unclear. In the present study, the root hydraulic properties of maize seedlings in response to pyrabactin were compared to those caused by ABA. Similar to ABA, lower concentration of pyrabactin induced a remarkable increase in Lpr as well as in the gene expression of the plasma membrane intrinsic protein (ZmPIP) aquaporin and in the ZmPIP2; 1/2; 2 protein abundance. The pyrabactin-induced enhancement of Lpr was abolished by H2O2 application, indicating that pyrabactin regulates Lpr by modulating ZmPIP at transcriptional, translational and post-translational (activity) level. Pyrabactin-mediated water transport and ZmPIP gene expression were phosphorylation-dependent, suggesting that ABA-PYR1-(PP2C)-protein kinase-AQP signaling pathway may be involved in this process. As we know this is the first established ABA signaling transduction pathway that mediated water transport in roots. This observation further addressed the importance of PYR1/PYLs ABA receptor in regulating plant water use efficiency from the under ground level. Except inhibiting transpiration in leaves, our result introduces the exciting possibility of application ABA agonists for regulating roots water uptake in field, with a species- and dose dependent manner.