ABSTRACT
Plants have their roots fixed in the soil, so they are unable to escape from adverse environments [...].
Subject(s)
Gene Expression Regulation , Plants , Plants/genetics , Soil , Plant Roots/geneticsABSTRACT
Plants respond to drought stress by producing abscisic acid, a chemical messenger that regulates gene expression and thereby expedites various physiological and cellular processes including the stomatal operation to mitigate stress and promote tolerance. To trigger or suppress gene transcription under drought stress conditions, the surrounding chromatin architecture must be converted between a repressive and active state by epigenetic remodeling, which is achieved by the dynamic interplay among DNA methylation, histone modifications, loop formation, and non-coding RNA generation. Plants can memorize chromatin status under drought conditions to enable them to deal with recurrent stress. Furthermore, drought tolerance acquired during plant growth can be transmitted to the next generation. The epigenetically modified chromatin architectures of memory genes under stressful conditions can be transmitted to newly developed cells by mitotic cell division, and to germline cells of offspring by overcoming the restraints on meiosis. In mammalian cells, the acquired memory state is completely erased and reset during meiosis. The mechanism by which plant cells overcome this resetting during meiosis to transmit memory is unclear. In this article, we review recent findings on the mechanism underlying transcriptional stress memory and the transgenerational inheritance of drought tolerance in plants.
Subject(s)
Droughts , Stress, Physiological , Animals , Stress, Physiological/genetics , Inheritance Patterns , Plants/genetics , Chromatin/genetics , Gene Expression Regulation, Plant , Plants, Genetically Modified/genetics , Mammals/geneticsABSTRACT
The current global climate crisis has led to drought, high salinity, and abnormaltemperatures (heat and cold), and is a serious threat to crop productivity. [...].
Subject(s)
Droughts , Stress, Physiological , Epigenesis, Genetic , Plants/genetics , SalinityABSTRACT
The plant hormone abscisic acid (ABA) triggers cellular tolerance responses to osmotic stress caused by drought and salinity. ABA controls the turgor pressure of guard cells in the plant epidermis, leading to stomatal closure to minimize water loss. However, stomatal apertures open to uptake CO2 for photosynthesis even under stress conditions. ABA modulates its signaling pathway via negative feedback regulation to maintain plant homeostasis. In the nuclei of guard cells, the clade A type 2C protein phosphatases (PP2Cs) counteract SnRK2 kinases by physical interaction, and thereby inhibit activation of the transcription factors that mediate ABA-responsive gene expression. Under osmotic stress conditions, PP2Cs bind to soluble ABA receptors to capture ABA and release active SnRK2s. Thus, PP2Cs function as a switch at the center of the ABA signaling network. ABA induces the expression of genes encoding repressors or activators of PP2C gene transcription. These regulators mediate the conversion of PP2C chromatins from a repressive to an active state for gene transcription. The stress-induced chromatin remodeling states of ABA-responsive genes could be memorized and transmitted to plant progeny; i.e., transgenerational epigenetic inheritance. This review focuses on the mechanism by which PP2C gene transcription modulates ABA signaling.
Subject(s)
Abscisic Acid/metabolism , Protein Phosphatase 2C/metabolism , Arabidopsis/genetics , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Chromatin Assembly and Disassembly/genetics , Chromatin Assembly and Disassembly/physiology , Droughts , Gene Expression Regulation, Plant/physiology , Osmotic Pressure/physiology , Signal Transduction/physiologyABSTRACT
Plants remember what they have experienced and are thereby able to confront repeated stresses more promptly and strongly. A subset of the drought responsive genes, called stress memory genes, displayed greatly elevated levels under recurrent drought conditions. To screen for a set of drought stress memory genes in soybean (Glycine max L.), we designed a 180K DNA chip comprising 60-bp probes synthesized in situ to examine 55,589 loci. Through microarray analysis using the DNA chip, we identified 2,162 and 2,385 genes with more than fourfold increases or decreases in transcript levels, respectively, under initial (first) drought stress conditions, when compared with the non-treated control. The transcript levels of the drought-responsive genes returned to basal levels during recovery (watered) states, and 392 and 613 genes displayed more than fourfold elevated or reduced levels, respectively, under subsequent (second) drought conditions, when compared to those observed under the first drought stress conditions. Gene Ontology and MapMan analyses classified the drought-induced memory genes exhibiting elevated levels of transcripts into several functional categories, including those involved in tolerance responses to abiotic stresses, which encode transcription factors, protein phosphatase 2Cs, and late embryogenesis abundant proteins. The drought-repressed memory genes exhibiting reduced levels of transcripts were classified into categories including photosynthesis and primary metabolism. Co-expression network analysis revealed that the soybean drought-induced and -repressed memory genes were equivalent to 172 and 311 Arabidopsis genes, respectively. The soybean drought stress memory genes include genes involved in the dehydration memory responses of Arabidopsis.
ABSTRACT
Type 2C protein phosphatases (PP2Cs) counteract protein kinases, thereby inhibiting the abscisic acid (ABA)-mediated response to abiotic stress in Arabidopsis thaliana. In the absence of stress, the promoters of PP2C genes (e.g., ABI1, ABI2, and HAI1) are negatively regulated by repressors that suppress gene transcription in a signal-independent manner. Quantitative reverse transcription PCR (RT-qPCR) and chromatin immunoprecipitation (ChIP) assays revealed that the levels of PP2C gene transcripts and RNA polymerase II (RNAPII) that stalled at the transcription start sites (TSS) of PP2C gene loci were increased under salt stress. The salt-induced increases in RNA polymerase-mediated transcription were reduced in 35S:AtMYB44 plants, confirming that AtMYB44 acts as a repressor of PP2C gene transcription. ChIP assays revealed that AtMYB44 repressors are released and nucleosomes are evicted from the promoter regions in response to salt stress. Under these conditions, histone H3 acetylation (H3ac) and methylation (H3K4me3) around the TSS regions significantly increased. The salt-induced increases in PP2C gene transcription were reduced in abf3 plants, indicating that ABF3 activates PP2C gene transcription. Overall, our data indicate that salt stress converts PP2C gene chromatin from a repressor-associated suppression status to an activator-mediated transcription status. In addition, we observed that the Arabidopsis mutant brm-3, which is moderately defective in SWI2/SNF2 chromatin remodeling ATPase BRAHMA (BRM) activity, produced more PP2C gene transcripts under salt stress conditions, indicating that BRM ATPase contributes to the repression of PP2C gene transcription.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/genetics , Chromatin/chemistry , Nucleosomes/metabolism , Phosphoprotein Phosphatases/metabolism , Salt Stress , Adenosine Triphosphate/chemistry , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Chromatin Assembly and Disassembly , DNA Methylation , DNA-Directed RNA Polymerases/metabolism , Gene Expression Profiling , Gene Expression Regulation, Plant , Plants, Genetically Modified/genetics , Promoter Regions, Genetic , Stress, Physiological , Transcription Factors/genetics , Transcription Factors/metabolism , Transcription, GeneticABSTRACT
AtLEA4-5 is a member of the group 4 late embryogenesis abundant (LEA) proteins, which are involved in the tolerance of water deficit in Arabidopsis thaliana. Chromatin immunoprecipitation assays revealed that the transcription factor AtMYB44 bound directly to the AtLEA4-5 gene promoter region under normal conditions, but was eliminated in response to osmotic stress (mannitol treatment). A quantitative reverse transcription PCR assay revealed that transcription of the AtLEA4-5 gene was induced in response to either salt (salinity) or mannitol (osmosis) treatment. The abiotic stress-induced increase in AtLEA4-5 transcripts was reduced in 35S:AtMYB44 transgenic plants, indicating that the transcription factor AtMYB44 represses gene transcription. More RNA polymerase II stalled at the transcription start site (TSS) of the AtLEA4-5 gene loci under osmotic stress, but the increment was reduced in the 35S:AtMYB44 plants. Histones are evicted from the promoter region under osmotic stress; however, histone eviction was hampered in the 35S:AtMYB44 plants. Under osmotic stress, the acetylated histones remaining at the TSS region was significantly lower in the 35S:AtMYB44 plants compared with wild-type plants. These results indicate that AtMYB44 suppresses polymerase-mediated transcription of the AtLEA4-5.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/genetics , Gene Expression Regulation, Plant , Molecular Chaperones/genetics , Transcription Factors/metabolism , Acetylation , Arabidopsis/physiology , Arabidopsis Proteins/genetics , Histones/metabolism , Osmoregulation , Osmotic Pressure , Promoter Regions, Genetic , Transcription, GeneticABSTRACT
AtMYB44 has been described in diverse hormonal signaling processes including abscisic acid (ABA)-mediated tolerance to abiotic stress; however, its function as a transcription factor is controversial. AtMYB44 contains the amino acid sequence LSLSL, a putative ETHYLENE-RESPONSIVE ELEMENT BINDING FACTOR-ASSOCIATED AMPHIPHILIC REPRESSION (EAR) motif. In yeast two-hybrid assay, physical interaction between AtMYB44 and a TOPLESS-RELATED (TPR) corepressor was observed, but abolished by mutation of the EAR motif. We performed bimolecular fluorescence complementation assay to confirm their interaction in planta. Chromatin immunoprecipitation assay revealed binding of AtMYB44 to the promoter regions of clade A protein phosphatase 2C (PP2C) genes (e.g., ABI1, ABI2, and HAI1), implying putative targets. Levels of histone H3 acetylation around the promoter regions were markedly lower in AtMYB44-overexpressing (35S:AtMYB44) plants than in wild-type plants. These results suggest that AtMYB44 forms a complex with TPR corepressors and recruits histone deacetylase(s) to suppress PP2C gene transcription in a signal-independent manner.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/genetics , Co-Repressor Proteins/metabolism , Gene Expression Regulation, Plant , Transcription Factors/metabolism , Transcription, Genetic , Acetylation , Genetic Loci , Histones/metabolism , Plants, Genetically Modified , Promoter Regions, Genetic/genetics , Protein Binding , Two-Hybrid System TechniquesABSTRACT
Plants have evolved strategies to cope with drought stress by maximizing physiological capacity and adjusting developmental processes such as flowering time. The WOX13 orthologous group is the most conserved among the clade of WOX homeodomain-containing proteins and is found to function in both drought stress and flower development. In this study, we isolated and characterized OsWOX13 from rice. OsWOX13 was regulated spatially in vegetative organs but temporally in flowers and seeds. Overexpression of OsWOX13 (OsWOX13-ov) in rice under the rab21 promoter resulted in drought resistance and early flowering by 7-10 days. Screening of gene expression profiles in mature leaf and panicles of OsWOX13-ov showed a broad spectrum of effects on biological processes, such as abiotic and biotic stresses, exerting a cross-talk between responses. Protein binding microarray and electrophoretic mobility shift assay analyses supported ATTGATTG as the putative cis-element binding of OsWOX13. OsDREB1A and OsDREB1F, drought stress response transcription factors, contain ATTGATTG motif(s) in their promoters and are preferentially expressed in OsWOX13-ov. In addition, Heading date 3a and OsMADS14, regulators in the flowering pathway and development, were enhanced in OsWOX13-ov. These results suggest that OsWOX13 mediates the stress response and early flowering and, thus, may be a regulator of genes involved in drought escape.
Subject(s)
Homeodomain Proteins/genetics , Oryza/physiology , Plant Proteins/genetics , Transcription Factors/genetics , Droughts , Flowers/genetics , Flowers/metabolism , Gene Expression Regulation, Plant , Genes, Homeobox , Homeodomain Proteins/metabolism , Oryza/genetics , Oryza/metabolism , Plant Proteins/metabolism , Stress, Physiological/genetics , Transcription Factors/metabolismABSTRACT
AtMYB44 transcripts accumulate non-specifically under diverse stress conditions and with various phytohormone treatments in Arabidopsis thaliana. We investigated the chromatin modifications caused by various signals to uncover the induction mechanism of AtMYB44 transcription. Bisulfite sequencing confirmed a previous database illustrating that the AtMYB44 promoter and gene-body regions are completely DNA methylation-free. Chromatin immunoprecipitation (ChIP) assays revealed that the nucleosome density is remarkably low at the AtMYB44 promoter region. Thus, the promoter region appears to be highly accessible for various trans-acting factors. ChIP assays revealed that osmotic stress (mannitol treatment) lowered the nucleosome density at the gene-body regions, while abscisic acid (ABA) or jasmonic acid (JA) treatment did so at the proximal transcription start site (TSS) region. In response to mannitol treatment, histone H3 lysine 4 trimethylation (H3K4me3) and H3 acetylation (H3ac) levels within the promoter, TSS, and gene-body regions of AtMYB44 were significantly increased. However, occupancy of histone variant H2A.Z was not affected by the mannitol treatment. We previously reported that salt stress triggered a significant decrease in H2A.Z occupation without affecting the H3K4me3 and H3ac levels. In combination, our data suggest that each signal transduced to the highly accessible promoter induces a different chromatin modification for AtMYB44 transcription.
Subject(s)
Arabidopsis Proteins/metabolism , Chromatin/metabolism , Gene Expression Regulation, Plant/drug effects , Transcription Factors/metabolism , Abscisic Acid/pharmacology , Arabidopsis , Arabidopsis Proteins/genetics , Chromatin Immunoprecipitation , Cyclopentanes/pharmacology , DNA Methylation , Mannitol/pharmacology , Mutation , Nucleosomes , Osmotic Pressure , Oxylipins/pharmacology , Promoter Regions, Genetic , Signal Transduction , Sodium Chloride/pharmacology , Transcription Factors/genetics , Transcription, Genetic , WaterABSTRACT
Transcripts of the Arabidopsis transcription factor gene, AtMYB44, accumulate rapidly to mediate a tolerance mechanism in response to salt stress. The AtMYB44 promoter is activated by salt stress, as illustrated in AtMYB44pro::GUS transgenic plants. Chromatin immunoprecipitation (ChIP) assays revealed that RNA polymerases were enriched on the AtMYB44 gene, especially on TSS-proximal regions, and nucleosome density was markedly reduced in the AtMYB44 gene-body region in response to salt stress. In addition, H2A.Z occupation was significantly decreased at the AtMYB44 promoter, transcription start site (TSS), and gene-body regions. Histone modifications including histone H3 lysine 4 trimethylation (H3K4me3) and histone H3 and H4 acetylation (H3ac and H4ac) were not affected under the same stress conditions. We found a decrease in the number of AtMYB44 proteins bound to their own gene promoters in response to salt stress. These results suggest that salt stress induces the eviction of H2A.Z-containing nucleosomes from the AtMYB44 promoter region, which may weaken its affinity for binding AtMYB44 protein that acts as a repressor for AtMYB44 gene transcription under salt stress-free conditions.
Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Arabidopsis/genetics , Arabidopsis/physiology , Histones/metabolism , Nucleosomes/metabolism , Sodium Chloride/pharmacology , Stress, Physiological/genetics , Transcription Factors/genetics , Transcription, Genetic , Arabidopsis/drug effects , Gene Expression Regulation, Plant/drug effects , Plants, Genetically Modified , Promoter Regions, Genetic , Protein Binding/drug effects , Transcription Factors/metabolism , Transcription, Genetic/drug effectsABSTRACT
Transgenic overexpression of the Arabidopsis gene for jasmonic acid carboxyl methyltransferase (AtJMT) is involved in regulating jasmonate-related plant responses. To examine its role in the compositional profile of soybean (Glycine max), we compared the seeds from field-grown plants that over-express AtJMT with those of the non-transgenic, wild-type (WT) counterpart. Our analysis of chemical compositions included proximates, amino acids, fatty acids, isoflavones, and antinutrients. Overexpression of AtJMT in the seeds resulted in decreased amounts of tryptophan, palmitic acid, linolenic acid, and stachyose, but increased levels of gadoleic acid and genistein. In particular, seeds from the transgenic soybeans contained 120.0-130.5% more genistein and 60.5-82.1% less stachyose than the WT. A separate evaluation of ingredient values showed that all were within the reference ranges reported for commercially available soybeans, thereby demonstrating the substantial equivalence of these transgenic and non-transgenic seeds.
Subject(s)
Glycine max/chemistry , Methyltransferases/chemistry , Methyltransferases/genetics , Plants, Genetically Modified/chemistry , Seeds/chemistryABSTRACT
Plant abiotic stress tolerance has been modulated by engineering the trehalose synthesis pathway. However, many stress-tolerant plants that have been genetically engineered for the trehalose synthesis pathway also show abnormal development. The metabolic intermediate trehalose 6-phosphate has the potential to cause aberrations in growth. To avoid growth inhibition by trehalose 6-phosphate, we used a gene that encodes a bifunctional in-frame fusion (BvMTSH) of maltooligosyltrehalose synthase (BvMTS) and maltooligosyltrehalose trehalohydrolase (BvMTH) from the nonpathogenic bacterium Brevibacterium helvolum. BvMTS converts maltooligosaccharides into maltooligosyltrehalose and BvMTH releases trehalose. Transgenic rice plants that over-express BvMTSH under the control of the constitutive rice cytochrome c promoter (101MTSH) or the ABA-inducible Ai promoter (105MTSH) show enhanced drought tolerance without growth inhibition. Moreover, 101MTSH and 105MTSH showed an ABA-hyposensitive phenotype in the roots. Our results suggest that over-expression of BvMTSH enhances drought-stress tolerance without any abnormal growth and showes ABA hyposensitive phenotype in the roots.
Subject(s)
Bacterial Proteins , Brevibacterium/enzymology , Droughts , Glucosidases , Glucosyltransferases , Abscisic Acid/pharmacology , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Gene Expression , Glucosidases/genetics , Glucosidases/metabolism , Glucosyltransferases/genetics , Glucosyltransferases/metabolism , Oligosaccharides/metabolism , Oryza/drug effects , Oryza/growth & development , Oryza/metabolism , Phenotype , Plant Growth Regulators/pharmacology , Plant Proteins/genetics , Plant Roots/drug effects , Plant Roots/metabolism , Plants, Genetically Modified/growth & development , Plants, Genetically Modified/metabolism , Promoter Regions, Genetic , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Sugar Phosphates/metabolism , Trehalose/analogs & derivatives , Trehalose/metabolismABSTRACT
AtMYB44 is a member of the R2R3 MYB subgroup 22 transcription factors and regulates diverse cellular responses in Arabidopsis thaliana. We performed quadruple 9-merbased protein binding microarray (PBM) analysis, which revealed that full-size AtMYB44 recognized and bound to the consensus sequence AACnG, where n represents A, G, C or T. The consensus sequence was confirmed by electrophoretic mobility shift assay (EMSA) with a truncated AtMYB44 protein containing the N-terminal side R2R3 domain. This result indicates that the R2R3 domain alone is sufficient to exhibit AtMYB44 binding specificity. The sequence AACnG is the type I binding site for MYB transcription factors, including all members of the subgroup 22. EMSA showed that the R2R3 domain protein binds in vitro to promoters of randomly selected Arabidopsis genes that contain the consensus binding sequence. This implies that AtMYB44 binds to any promoter region that contains the consensus sequence, without determining their functional activity or specificity. The C-terminal side transcriptional activation domain of AtMYB44 contains an asparagine-rich fragment, NINNTTSSRHNHNN (aa 215-228), which, among the members of subgroup 22, is unique to AtMYB44. A transcriptional activation assay in yeast showed that this fragment is included in a region (aa 200-240) critical for the ability of AtMYB44 to function as a transcriptional activator. We hypothesize that the C-terminal side of the protein, but not the N-terminal side of the R2R3 domain, contributes to the functional activity and specificity of AtMYB44 through interactions with other regulators generated by each of a variety of stimuli.
Subject(s)
Arabidopsis Proteins/genetics , Regulatory Sequences, Nucleic Acid , Transcription Factors/genetics , Amino Acid Sequence , Arabidopsis Proteins/metabolism , Base Sequence , Binding Sites , Electrophoretic Mobility Shift Assay , Microarray Analysis , Molecular Sequence Data , Promoter Regions, Genetic , Protein Binding , Sequence Alignment , Transcription Factors/metabolismABSTRACT
Jasmonates play important roles in development, stress responses and defense in plants. Here, we report the results of a study using a functional genomics approach that identified a rice basic helix-loop-helix domain gene, OsbHLH148, that conferred drought tolerance as a component of the jasmonate signaling module in rice. OsbHLH148 transcript levels were rapidly increased by treatment with methyl jasmonate (MeJA) or abscisic acid, and abiotic stresses including dehydration, high salinity, low temperature and wounding. Transgenic over-expression of OsbHLH148 in rice confers plant tolerance to drought stress. Expression profiling followed by DNA microarray and RNA gel-blot analyses of transgenic versus wild-type rice identified genes that are up-regulated by OsbHLH148 over-expression. These include OsDREB and OsJAZ genes that are involved in stress responses and the jasmonate signaling pathway, respectively. OsJAZ1, a rice ZIM domain protein, interacted with OsbHLH148 in yeast two-hybrid and pull-down assays, but it interacted with the putative OsCOI1 only in the presence of coronatine. Furthermore, the OsJAZ1 protein was degraded by rice and Arabidopsis extracts in the presence of coronatine, and its degradation was inhibited by MG132, a 26S proteasome inhibitor, suggesting 26S proteasome-mediated degradation of OsJAZ1 via the SCF(OsCOI1) complex. The transcription level of OsJAZ1 increased upon exposure of rice to MeJA. These results show that OsJAZ1 could act as a transcriptional regulator of the OsbHLH148-related jasmonate signaling pathway leading to drought tolerance. Thus, our study suggests that OsbHLH148 acts on an initial response of jasmonate-regulated gene expression toward drought tolerance, constituting the OsbHLH148-OsJAZ-OsCOI1 signaling module in rice.
Subject(s)
Basic Helix-Loop-Helix Transcription Factors/genetics , Basic Helix-Loop-Helix Transcription Factors/metabolism , Cyclopentanes/metabolism , Oryza/genetics , Oryza/metabolism , Oxylipins/metabolism , Plant Proteins/genetics , Plant Proteins/metabolism , Amino Acid Sequence , Base Sequence , DNA, Plant/genetics , Droughts , Gene Expression Profiling , Gene Expression Regulation, Plant , Genes, Plant , Models, Biological , Molecular Sequence Data , Oligonucleotide Array Sequence Analysis , Plants, Genetically Modified , Sequence Homology, Amino Acid , Signal Transduction , Stress, Physiological , Up-RegulationABSTRACT
Arabidopsis RNA polymerase II (RNAPII) C-terminal domain (CTD) phosphatases regulate stress-responsive gene expression and plant development via the dephosphorylation of serine (Ser) residues of the CTD. Some of these phosphatases (CTD phosphatase-like 1 (CPL1) to CPL3) negatively regulate ABA and stress responses. Here, we isolated AtCPL5, a cDNA encoding a protein containing two CTD phosphatase domains (CPDs). To characterize AtCPL5, we analyzed the gene expression patterns and subcellular protein localization, investigated various phenotypes of AtCPL5-overexpressors and knockout mutants involved in ABA and drought responses, performed microarray and RNA hybridization analyses using AtCPL5-overexpressors, and assessed the CTD phosphatase activities of the purified AtCPL5 and each CPD of the protein. Transcripts of the nucleus-localized AtCPL5 were induced by ABA and drought. AtCPL5-overexpressors exhibited ABA-hypersensitive phenotypes (increased inhibition of seed germination, seedling growth, and stomatal aperture), lower transpiration rates upon dehydration, and enhanced drought tolerance, while the knockout mutants showed weak ABA hyposensitivity. AtCPL5 overexpression changed the expression of numerous genes, including those involved in ABA-mediated responses. In contrast to Ser-5-specific phosphatase activity of the negative stress response regulators, purified AtCPL5 and each CPD of the protein specifically dephosphorylated Ser-2 in RNAPII CTD. We conclude that AtCPL5 is a unique CPL family protein that positively regulates ABA-mediated development and drought responses in Arabidopsis.
Subject(s)
Abscisic Acid/pharmacology , Arabidopsis Proteins/metabolism , Arabidopsis/enzymology , Arabidopsis/physiology , Droughts , Phosphoserine/metabolism , Amino Acid Sequence , Arabidopsis/drug effects , Arabidopsis/genetics , Arabidopsis Proteins/chemistry , Arabidopsis Proteins/genetics , Cell Nucleus/drug effects , Cell Nucleus/metabolism , Cloning, Molecular , DNA, Complementary/genetics , Gene Expression Regulation, Plant/drug effects , Gene Knockout Techniques , Genes, Plant , Glucuronidase/metabolism , Molecular Sequence Data , Phylogeny , Plant Stomata/drug effects , Plants, Genetically Modified , Protein Structure, Tertiary , RNA Polymerase II/chemistry , RNA Polymerase II/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Real-Time Polymerase Chain Reaction , Stress, Physiological/drug effects , Stress, Physiological/genetics , Subcellular Fractions/drug effects , Subcellular Fractions/metabolism , Up-Regulation/drug effects , Up-Regulation/geneticsABSTRACT
The Arabidopsis thaliana transcription factor gene AtMYB44 was induced within 10 min by treatment with methyl jasmonate (MeJA). Wound-induced expression of the gene was observed in local leaves, but not in distal leaves, illustrating jasmonate-independent induction at wound sites. AtMYB44 expression was not abolished in Arabidopsis mutants insensitive to jasmonate (coi1), ethylene (etr1), or abscisic acid (abi3-1) when treated with the corresponding hormones. Moreover, various growth hormones and sugars also induced rapid AtMYB44 transcript accumulation. Thus, AtMYB44 gene activation appears to not be induced by any specific hormone. MeJA-induced activation of jasmonate-responsive genes such as JR2, VSP, LOXII, and AOS was attenuated in transgenic Arabidopsis plants overexpressing the gene (35S:AtMYB44), but significantly enhanced in atmyb44 knockout mutants. The 35S:MYB44 and atmyb44 plants did not show defectiveness in MeJA-induced primary root growth inhibition, indicating that the differences in jasmonate-responsive gene expression observed was not due to alterations in the jasmonate signaling pathway. 35S:AtMYB44 seedlings exhibited slightly elevated chlorophyll levels and less jasmonate- induced anthocyanin accumulation, demonstrating suppression of jasmonate-mediated responses and enhancement of ABA-mediated responses. These observations support the hypothesis of mutual antagonistic actions between jasmonate- and abscisic acid-mediated signaling pathways.
Subject(s)
Abscisic Acid/metabolism , Acetates/metabolism , Arabidopsis Proteins/biosynthesis , Arabidopsis/genetics , Cyclopentanes/metabolism , Oxylipins/metabolism , Plant Growth Regulators/metabolism , Transcription Factors/biosynthesis , Anthocyanins/metabolism , Arabidopsis/growth & development , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Chlorophyll/metabolism , DNA-Binding Proteins/biosynthesis , DNA-Binding Proteins/genetics , Ethylenes/metabolism , Gene Expression Regulation, Plant , Genes, abl/genetics , Plants, Genetically Modified , Receptors, Cell Surface/biosynthesis , Receptors, Cell Surface/genetics , Signal Transduction , Transcription Factors/genetics , Transcriptional ActivationABSTRACT
The Arabidopsis gene AtLEC (At3g15356) gene encodes a putative 30-kDa protein with a legume lectin-like domain. Likely to classic legume lectin family of genes, AtLEC is expressed in rosette leaves, primary inflorescences, and roots, as observed in Northern blot analysis. The accumulation of AtLEC transcript is induced very rapidly, within 30 min, by chitin, a fungal wall-derived oligosaccharide elictor of the plant defense response. Transgenic Arabidopsis carrying an AtLEC promoter-driven beta-glucuronidase (GUS) construct exhibited GUS activity in the leaf veins, secondary inflorescences, carpel heads, and silique receptacles, in which no expression could be seen in Northern blot analysis. This observation suggests that AtLEC expression is induced transiently and locally during developmental processes in the absence of an external signal such as chitin. In addition, mechanically wounded sites showed strong GUS activity, indicating that the AtLEC promoter responds to jasmonate. Indeed, methyl jasmonate and ethylene exposure induced AtLEC expression within 3-6 h. Thus, the gene appears to play a role in the jasmonate-/ethylene-responsive, in addition to the chitin-elicited, defense responses. However, chitin-induced AtLEC expression was also observed in jasmonate-insensitive (coi1) and ethylene-insensitive (etr1-1) Arabidopsis mutants. Thus, it appears that chitin promotes AtLEC expression via a jasmonate- and/or ethylene-independent pathway.
Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis/growth & development , Arabidopsis/genetics , Chitin/pharmacology , Plant Growth Regulators/pharmacology , Plant Lectins/genetics , Up-Regulation/drug effects , Acetates/pharmacology , Amino Acid Sequence , Arabidopsis/drug effects , Arabidopsis Proteins/chemistry , Blotting, Northern , Cyclopentanes/pharmacology , Ethylenes/pharmacology , Gene Expression Profiling , Gene Expression Regulation, Plant/drug effects , Genes, Plant , Glucuronidase/metabolism , Molecular Sequence Data , Organ Specificity/drug effects , Oxylipins/pharmacology , Plant Lectins/chemistry , Signal Transduction/drug effectsABSTRACT
MhMTS and MhMTH are trehalose (alpha-D-glucopyranosyl- [1,1]-alpha-D-glucopyranose) biosynthesis genes of the thermophilic microorganism Metallosphaera hakonensis, and encode a maltooligosyltrehalose synthase (MhMTS) and a maltooligosyltrehalose trehalohydrolase (MhMTH), respectively. In this study, the two genes were fused inframe in a recombinant DNA, and expressed in Escherichia coli to produce a bifunctional fusion enzyme, MhMTSH. Similar to the two-step reactions with MhMTS and MhMTH, the fusion enzyme catalyzed the sequential reactions on maltopentaose, maltotriosyltrehalose formation, and following hydrolysis, producing trehalose and maltotriose. Optimum conditions for the fusion enzyme-catalyzed trehalose synthesis were around 70 degrees and pH 5.0-6.0. The MhMTSH fusion enzyme exhibited a high degree of thermostability, retaining 80% of the activity when pre-incubated at 70 degrees for 48 h. The stability was gradually abolished by incubating the fusion enzyme at above 80 degrees . The MhMTSH fusion enzyme was active on various sizes of maltooligosaccharides, extending its substrate specificity to soluble starch, the most abundant natural source of trehalose production.
Subject(s)
Glucosidases/metabolism , Glucosyltransferases/metabolism , Sulfolobaceae/enzymology , Trehalose/biosynthesis , Chromatography, Ion Exchange , Chromatography, Thin Layer , Cloning, Molecular , Escherichia coli/genetics , Glucosidases/genetics , Glucosyltransferases/genetics , Hot Temperature , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Starch/metabolism , Sulfolobaceae/geneticsABSTRACT
We developed a quantitative method for the determination of methyl esterase activity, analyzing substrate specificity against three major signal molecules, jasmonic acid methyl ester (MeJA), salicylic acid methyl ester (MeSA), and indole-3-acetic acid methyl ester (MeIAA). We used a silylation reagent for chemical derivatization and used gas chromatography (GC)-mass spectroscopy in analyses, for high precision. To test this method, an Arabidopsis esterase gene, AtME8, was expressed in Escherichia coli, and then the kinetic parameters of the recombinant enzyme were determined for three substrates. Finally, this method was also applied to the direct quantification of phytohormones in petals from lilies and roses.