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1.
Front Plant Sci ; 12: 751852, 2021.
Article in English | MEDLINE | ID: mdl-34707632

ABSTRACT

Stomata are micropores that allow plants to breathe and play a critical role in photosynthesis and nutrient uptake by regulating gas exchange and transpiration. Stomatal development, therefore, is optimized for survival and growth of the plant despite variable environmental conditions. Signaling cascades and transcriptional networks that determine the birth, proliferation, and differentiation of a stomate have been identified. These networks ensure proper stomatal patterning, density, and polarity. Environmental cues also influence stomatal development. In this review, we highlight recent findings regarding the developmental program governing cell fate and dynamics of stomatal lineage cells at the cell state- or single-cell level. We also overview the control of stomatal development by environmental cues as well as developmental plasticity associated with stomatal function and physiology. Recent advances in our understanding of stomatal development will provide a route to improving photosynthesis and water-stress resilience of crop plants in the climate change we currently face.

2.
Front Plant Sci ; 11: 551, 2020.
Article in English | MEDLINE | ID: mdl-32499801

ABSTRACT

Leaf senescence is a developmental process designed for nutrient recycling and relocation to maximize growth competence and reproductive capacity of plants. Thus, plants integrate developmental and environmental signals to precisely control senescence. To genetically dissect the complex regulatory mechanism underlying leaf senescence, we identified an early leaf senescence mutant, rse1. RSE1 encodes a putative glycosyltransferase. Loss-of-function mutations in RSE1 resulted in precocious leaf yellowing and up-regulation of senescence marker genes, indicating enhanced leaf senescence. Transcriptome analysis revealed that salicylic acid (SA) and defense signaling cascades were up-regulated in rse1 prior to the onset of leaf senescence. We found that SA accumulation was significantly increased in rse1. The rse1 phenotypes are dependent on SA-INDUCTION DEFICIENT 2 (SID2), supporting a role of SA in accelerated leaf senescence in rse1. Furthermore, RSE1 protein was localized to the cell wall, implying a possible link between the cell wall and RSE1 function. Together, we show that RSE1 negatively modulates leaf senescence through an SID2-dependent SA signaling pathway.

3.
Proc Natl Acad Sci U S A ; 117(11): 6237-6245, 2020 03 17.
Article in English | MEDLINE | ID: mdl-32123075

ABSTRACT

Stomata in the plant epidermis play a critical role in growth and survival by controlling gas exchange, transpiration, and immunity to pathogens. Plants modulate stomatal cell fate and patterning through key transcriptional factors and signaling pathways. MicroRNAs (miRNAs) are known to contribute to developmental plasticity in multicellular organisms; however, no miRNAs appear to target the known regulators of stomatal development. It remains unclear as to whether miRNAs are involved in stomatal development. Here, we report highly dynamic, developmentally stage-specific miRNA expression profiles from stomatal lineage cells. We demonstrate that stomatal lineage miRNAs positively and negatively regulate stomatal formation and patterning to avoid clustered stomata. Target prediction of stomatal lineage miRNAs implicates potential cellular processes in stomatal development. We show that miR399-mediated PHO2 regulation, involved in phosphate homeostasis, contributes to the control of stomatal development. Our study demonstrates that miRNAs constitute a critical component in the regulatory mechanisms controlling stomatal development.


Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis/physiology , Gene Expression Regulation, Plant/physiology , MicroRNAs/metabolism , Plant Stomata/growth & development , Ubiquitin-Conjugating Enzymes/genetics , MicroRNAs/genetics , Plants, Genetically Modified , RNA-Seq
4.
J Exp Bot ; 71(1): 178-187, 2020 01 01.
Article in English | MEDLINE | ID: mdl-31563952

ABSTRACT

Seed germination is a developmental process regulated by numerous internal and external cues. Our previous studies have shown that calcium influx mediated by the Arabidopsis glutamate receptor homolog 3.5 (AtGLR3.5) modulates the expression of the ABSCISIC ACID INSENSITIVE 4 (ABI4) transcription factor during germination and that L-methionine (L-Met) activates AtGLR3.1/3.5 Ca2+ channels in guard cells. However, it is not known whether L-Met participates in regulation of germination and what cellular mechanism is responsible for Met production during germination. Here, we describe Arabidopsis methionine synthase 1 (AtMS1), which acts in the final step of Met biosynthesis, synthesizes the Met required for the activation of AtGLR3.5 Ca2+ channels whose expression is up-regulated during germination, leading to the regulation of seed germination. We show that exogenous L-Met promotes germination in an AtGRL3.5-dependent manner. We also demonstrate that L-Met directly regulates the AtGLR3.5-mediated increase in cytosolic Ca2+ level in seedlings. We provide pharmacological and genetic evidence that Met synthesized via AtMS1 acts upstream of the AtGLR3.5-mediated Ca2+ signal and regulates the expression of ABI4, a major regulator in the abscisic acid response in seeds. Overall, our results link AtMS1, L-Met, the AtGLR3.5 Ca2+ channel, Ca2+ signals, and ABI4, and shed light on the physiological role and molecular mechanism of L-Met in germination.


Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis/physiology , Gene Expression Regulation, Plant , Germination/genetics , Methionine/metabolism , Receptors, Glutamate/genetics , Transcription Factors/genetics , Arabidopsis/genetics , Arabidopsis Proteins/metabolism , Receptors, Glutamate/metabolism , Transcription Factors/metabolism
5.
Nat Plants ; 5(12): 1273-1282, 2019 12.
Article in English | MEDLINE | ID: mdl-31740770

ABSTRACT

RNA quality control (RQC) and post-transcriptional gene silencing (PTGS) target and degrade aberrant endogenous RNAs and foreign RNAs, contributing to homeostasis of cellular RNAs. In plants, RQC and PTGS compete for foreign and selected endogenous RNAs; however, little is known about the mechanism interconnecting the two pathways. Using a reporter system designed for monitoring PTGS, we revealed that the 26S proteasome subunit RPT2a enhances transgene PTGS by promoting the accumulation of transgene-derived short interfering RNAs without affecting their biogenesis. RPT2a physically associated with a subset of RQC components and downregulated the protein level. Overexpression of the RQC components interfered with transgene silencing, and impairment of the RQC machinery reinforced transgene PTGS attenuated by rpt2a. Overall, we demonstrate that the 26S proteasome subunit RPT2a promotes PTGS by repressing the RQC machinery to control foreign RNAs.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/genetics , Proteasome Endopeptidase Complex/metabolism , RNA Interference , RNA, Plant/genetics , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Gene Expression Regulation, Plant , Proteasome Endopeptidase Complex/genetics , RNA, Plant/metabolism , Transgenes
6.
Front Plant Sci ; 9: 803, 2018.
Article in English | MEDLINE | ID: mdl-30013580

ABSTRACT

Cellular calcium acts as a second messenger and regulates diverse developmental events and stress responses. Cytosolic calcium has long been considered as an important regulator of senescence, however, the role of Ca2+ in plant senescence has remained elusive. Here we show that the Calmodulin 1 (CaM1) gene, which encodes Ca2+-binding protein calmodulin 1, positively regulates leaf senescence in Arabidopsis. Yellowing of leaves, accumulation of reactive oxygen species (ROS), and expression of the senescence-associated gene 12 (SAG12) were significantly enhanced in CaM1 overexpression plants. In contrast, abscisic acid (ABA)-triggered ROS production and stomatal closure were reduced in amiRNA-CaM1 plants. We found a positive-feedback regulation loop among three signaling components, CaM1, RPK1, and RbohF, which physically associate with each other. RPK1 positively regulates the expression of the CaM1 gene, and the CaM1 protein, in turn, up-regulates RbohF gene expression. Interestingly, the expression of CaM1 was down-regulated in rbohD, rbohF, and rbohD/F mutants. We show that CaM1 positively regulates ROS production, leaf senescence, and ABA response in Arabidopsis.

7.
BMB Rep ; 51(7): 317-318, 2018 Jul.
Article in English | MEDLINE | ID: mdl-29966583

ABSTRACT

Plants are unable to relocate themselves to a more favorable location and thus have to deal with developmental programs and environmental cues wherever they happen to be. It is yet largely unknown how plant cells coordinate cellular activities and architectures to accomplish developmental processes and respond to environmental changes. By identifying and establishing a new cellular model system, we have discovered that two neighboring cell types in the abscission zone (AZ) of Arabidopsis flowers coordinate their activities to ensure a precise "cut" through a highly restricted area of plant tissue to bring about organ separation. From this perspective, we further discuss the essence of cellular coordination in AZ, the key molecules controlling the organ separation, and relevant implications. [BMB Reports 2018; 51(7): 317-318].


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/physiology , Cell Wall/metabolism , Arabidopsis/genetics , Arabidopsis/metabolism , Flowers/metabolism , Lignin/metabolism , Reactive Oxygen Species/metabolism
8.
Cell ; 173(6): 1468-1480.e9, 2018 05 31.
Article in English | MEDLINE | ID: mdl-29731167

ABSTRACT

The cell wall, a defining feature of plants, provides a rigid structure critical for bonding cells together. To overcome this physical constraint, plants must process cell wall linkages during growth and development. However, little is known about the mechanism guiding cell-cell detachment and cell wall remodeling. Here, we identify two neighboring cell types in Arabidopsis that coordinate their activities to control cell wall processing, thereby ensuring precise abscission to discard organs. One cell type produces a honeycomb structure of lignin, which acts as a mechanical "brace" to localize cell wall breakdown and spatially limit abscising cells. The second cell type undergoes transdifferentiation into epidermal cells, forming protective cuticle, demonstrating de novo specification of epidermal cells, previously thought to be restricted to embryogenesis. Loss of the lignin brace leads to inadequate cuticle formation, resulting in surface barrier defects and susceptible to infection. Together, we show how plants precisely accomplish abscission.


Subject(s)
Arabidopsis/physiology , Cell Wall/metabolism , Lignin/metabolism , Arabidopsis Proteins/metabolism , Cell Differentiation , Cell Membrane/metabolism , Gene Expression Profiling , Gene Expression Regulation, Plant , Mutation , NADPH Oxidases/metabolism , Plants, Genetically Modified/physiology , Pseudomonas syringae , Surface Properties
9.
Cell Rep ; 21(12): 3373-3380, 2017 Dec 19.
Article in English | MEDLINE | ID: mdl-29262318

ABSTRACT

Reactive oxygen species (ROS) are inevitable by-products of aerobic metabolic processes, causing non-specific oxidative damage and also acting as second messengers. Superoxide is a short-lived ROS that functions in various cellular responses, including aging and cell death. However, it is unclear as to how superoxide brings about age-dependent cell death and senescence. Here, we show that the accumulation and signaling of superoxide are mediated by three Arabidopsis proteins-RPK1, CaM4, and RbohF-which trigger subsequent cellular events leading to age-dependent cell death. We demonstrate that the NADPH oxidase RbohF is responsible for RPK1-mediated transient accumulation of superoxide, SIRK kinase induction, and cell death, all of which are positively regulated by CaM4. RPK1 physically interacts with and phosphorylates CaM4, which, in turn, interacts with RbohF. Overall, we demonstrate how the protein trio governs the superoxide accumulation and signaling at the cell surface to control senescence and cell death.


Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Calmodulin/metabolism , Cell Death , Cellular Senescence , NADPH Oxidases/metabolism , Protein Kinases/genetics , Superoxides/metabolism , Arabidopsis , Calmodulin/genetics , NADPH Oxidases/genetics , Protein Binding , Protein Kinases/metabolism
10.
Biosci Biotechnol Biochem ; 81(7): 1394-1400, 2017 Jul.
Article in English | MEDLINE | ID: mdl-28387156

ABSTRACT

Salicylic acid (SA) induces stomatal closure sharing several components with abscisic acid (ABA) and methyl jasmonate (MeJA) signaling. We have previously shown that two guard cell-preferential mitogen-activated protein kinases (MAPKs), MPK9 and MPK12, positively regulate ABA signaling and MeJA signaling in Arabidopsis thaliana. In this study, we examined whether these two MAPKs are involved in SA-induced stomatal closure using genetic mutants and a pharmacological, MAPKK inhibitor. Salicylic acid induced stomatal closure in mpk9 and mpk12 single mutants but not in mpk9 mpk12 double mutants. The MAPKK inhibitor PD98059 inhibited SA-induced stomatal closure in wild-type plants. Salicylic acid induced extracellular reactive oxygen species (ROS) production, intracellular ROS accumulation, and cytosolic alkalization in the mpk9, mpk12, and mpk9 mpk12 mutants. Moreover, SA-activated S-type anion channels in guard cells of wild-type plants but not in guard cells of mpk9 mpk12 double mutants. These results imply that MPK9 and MPK12 are positive regulators of SA signaling in Arabidopsis guard cells.


Subject(s)
Arabidopsis/drug effects , Gene Expression Regulation, Plant , Plant Stomata/drug effects , Salicylic Acid/pharmacology , Abscisic Acid/metabolism , Abscisic Acid/pharmacology , Acetates/metabolism , Acetates/pharmacology , Arabidopsis/genetics , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Cyclopentanes/metabolism , Cyclopentanes/pharmacology , Flavonoids/pharmacology , Hydrogen-Ion Concentration , Membrane Potentials/drug effects , Mitogen-Activated Protein Kinases/genetics , Mitogen-Activated Protein Kinases/metabolism , Mutation , Oxylipins/metabolism , Oxylipins/pharmacology , Plant Growth Regulators/metabolism , Plant Growth Regulators/pharmacology , Plant Stomata/genetics , Plant Stomata/metabolism , Protein Kinase Inhibitors/pharmacology , Reactive Oxygen Species/agonists , Reactive Oxygen Species/metabolism , Salicylic Acid/metabolism , Signal Transduction , Voltage-Dependent Anion Channels/genetics , Voltage-Dependent Anion Channels/metabolism
11.
Proc Natl Acad Sci U S A ; 113(51): 14858-14863, 2016 12 20.
Article in English | MEDLINE | ID: mdl-27930340

ABSTRACT

Histone acetylation is a major epigenetic control mechanism that is tightly linked to the promotion of gene expression. Histone acetylation levels are balanced through the opposing activities of histone acetyltransferases (HATs) and histone deacetylases (HDACs). Arabidopsis HDAC genes (AtHDACs) compose a large gene family, and distinct phenotypes among AtHDAC mutants reflect the functional specificity of individual AtHDACs However, the mechanisms underlying this functional diversity are largely unknown. Here, we show that POWERDRESS (PWR), a SANT (SWI3/DAD2/N-CoR/TFIII-B) domain protein, interacts with HDA9 and promotes histone H3 deacetylation, possibly by facilitating HDA9 function at target regions. The developmental phenotypes of pwr and hda9 mutants were highly similar. Three lysine residues (K9, K14, and K27) of H3 retained hyperacetylation status in both pwr and hda9 mutants. Genome-wide H3K9 and H3K14 acetylation profiling revealed elevated acetylation at largely overlapping sets of target genes in the two mutants. Highly similar gene-expression profiles in the two mutants correlated with the histone H3 acetylation status in the pwr and hda9 mutants. In addition, PWR and HDA9 modulated flowering time by repressing AGAMOUS-LIKE 19 expression through histone H3 deacetylation in the same genetic pathway. Finally, PWR was shown to physically interact with HDA9, and its SANT2 domain, which is homologous to that of subunits in animal HDAC complexes, showed specific binding affinity to acetylated histone H3. We therefore propose that PWR acts as a subunit in a complex with HDA9 to result in lysine deacetylation of histone H3 at specific genomic targets.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/genetics , Histone Deacetylases/metabolism , Transcription Factors/metabolism , Acetylation , Arabidopsis/metabolism , Gene Expression Regulation, Plant , Histones/chemistry , Humans , Lysine/chemistry , Mutation , Protein Domains , Protein Processing, Post-Translational , Transcriptome
12.
Cell Rep ; 17(10): 2553-2561, 2016 12 06.
Article in English | MEDLINE | ID: mdl-27926860

ABSTRACT

Plant glutamate receptor homologs (GLRs) have long been proposed to function as ligand-gated Ca2+ channels, but no in planta evidence has been provided. Here, we present genetic evidence that Arabidopsis GLR3.1 and GLR3.5 form Ca2+ channels activated by L-methionine (L-Met) at physiological concentrations and regulate stomatal apertures and plant growth. The glr3.1/3.5 mutations resulted in a lower cytosolic Ca2+ level, defective Ca2+-induced stomatal closure, and Ca2+-deficient growth disorder, all of which involved L-Met. Patch-clamp analyses of guard cells showed that GLR3.1/3.5 Ca2+ channels are activated specifically by L-Met, with the activation abolished in glr3.1/3.5. Moreover, GLR3.1/3.5 Ca2+ channels are distinct from previously characterized ROS-activated Ca2+ channels and act upstream of ROS, providing Ca2+ transients necessary for the activation of NADPH oxidases. Our data indicate that GLR3.1/3.5 constitute L-Met-activated Ca2+ channels responsible for maintaining basal [Ca2+]cyt, play a pivotal role in plant growth, and act upstream of ROS, thereby regulating stomatal aperture.


Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis/genetics , Calcium/metabolism , Methionine/metabolism , Receptors, Glutamate/genetics , Arabidopsis/growth & development , Arabidopsis/metabolism , Arabidopsis Proteins/metabolism , Cytosol/metabolism , Mutation , NADPH Oxidases/metabolism , Nitric Oxide/metabolism , Reactive Oxygen Species/metabolism , Receptors, Glutamate/metabolism , Signal Transduction/genetics
13.
Front Plant Sci ; 7: 80, 2016.
Article in English | MEDLINE | ID: mdl-26904052

ABSTRACT

Guard cells form stomata on the epidermis and continuously respond to endogenous and environmental stimuli to fine-tune the gas exchange and transpirational water loss, processes which involve mitogen-activated protein kinase (MAPK) cascades. MAPKs form three-tiered kinase cascades with MAPK kinases and MAPK kinase kinases, by which signals are transduced to the target proteins. MAPK cascade genes are highly conserved in all eukaryotes, and they play crucial roles in myriad developmental and physiological processes. MAPK cascades function during biotic and abiotic stress responses by linking extracellular signals received by receptors to cytosolic events and gene expression. In this review, we highlight recent findings and insights into MAPK-mediated guard cell signaling, including the specificity of MAPK cascades and the remaining questions.

14.
New Phytol ; 210(4): 1169-89, 2016 06.
Article in English | MEDLINE | ID: mdl-26879345

ABSTRACT

1169 I. 1170 II. 1170 III. 1172 IV. 1176 V. 1181 VI. 1182 1183 References 1183 SUMMARY: Modern agriculture is facing multiple challenges including the necessity for a substantial increase in production to meet the needs of a burgeoning human population. Water shortage is a deleterious consequence of both population growth and climate change and is one of the most severe factors limiting global crop productivity. Brassica species, particularly canola varieties, are cultivated worldwide for edible oil, animal feed, and biodiesel, and suffer dramatic yield loss upon drought stress. The recent release of the Brassica napus genome supplies essential genetic information to facilitate identification of drought-related genes and provides new information for agricultural improvement in this species. Here we summarize current knowledge regarding drought responses of canola, including physiological and -omics effects of drought. We further discuss knowledge gained through translational biology based on discoveries in the closely related reference species Arabidopsis thaliana and through genetic strategies such as genome-wide association studies and analysis of natural variation. Knowledge of drought tolerance/resistance responses in canola together with research outcomes arising from new technologies and methodologies will inform novel strategies for improvement of drought tolerance and yield in this and other important crop species.


Subject(s)
Genome, Plant/genetics , Agriculture , Arabidopsis/genetics , Arabidopsis/physiology , Brassica napus/genetics , Brassica napus/physiology , Climate Change , Crops, Agricultural , Droughts , Genome-Wide Association Study , Stress, Physiological
15.
Mol Plant ; 9(3): 447-460, 2016 Mar 07.
Article in English | MEDLINE | ID: mdl-26724418

ABSTRACT

Stomatal movements are critical in regulating gas exchange for photosynthesis and water balance between plant tissues and the atmosphere. The plant hormone abscisic acid (ABA) plays key roles in regulating stomatal closure under various abiotic stresses. In this study, we revealed a novel role of BAK1 in guard cell ABA signaling. We found that the brassinosteroid (BR) signaling mutant bak1 lost more water than wild-type plants and showed ABA insensitivity in stomatal closure. ABA-induced OST1 expression and reactive oxygen species (ROS) production were also impaired in bak1. Unlike direct treatment with H2O2, overexpression of OST1 did not completely rescue the insensitivity of bak1 to ABA. We demonstrated that BAK1 forms a complex with OST1 near the plasma membrane and that the BAK1/OST1 complex is increased in response to ABA in planta. Brassinolide, the most active BR, exerted a negative effect on ABA-induced formation of the BAK1/OST1 complex and OST1 expression. Moreover, we found that BAK1 and ABI1 oppositely regulate OST1 phosphorylation in vitro, and that ABI1 interacts with BAK1 and inhibits the interaction of BAK1 and OST1. Taken together, our results suggest that BAK1 regulates ABA-induced stomatal closure in guard cells.


Subject(s)
Abscisic Acid/metabolism , Arabidopsis Proteins/metabolism , Arabidopsis/cytology , Arabidopsis/metabolism , Protein Kinases/metabolism , Signal Transduction , Abscisic Acid/pharmacology , Arabidopsis/anatomy & histology , Arabidopsis/drug effects , Arabidopsis Proteins/genetics , Brassinosteroids/pharmacology , Cell Membrane/drug effects , Cell Membrane/metabolism , Gene Expression Regulation, Plant/drug effects , Mutation , Phosphorylation/drug effects , Plant Stomata/anatomy & histology , Plant Stomata/drug effects , Protein Kinases/genetics , Reactive Oxygen Species/metabolism , Signal Transduction/drug effects , Steroids, Heterocyclic/pharmacology
16.
Plant Physiol ; 167(4): 1630-42, 2015 Apr.
Article in English | MEDLINE | ID: mdl-25681329

ABSTRACT

Seed germination is a critical step in a plant's life cycle that allows successful propagation and is therefore strictly controlled by endogenous and environmental signals. However, the molecular mechanisms underlying germination control remain elusive. Here, we report that the Arabidopsis (Arabidopsis thaliana) glutamate receptor homolog3.5 (AtGLR3.5) is predominantly expressed in germinating seeds and increases cytosolic Ca2+ concentration that counteracts the effect of abscisic acid (ABA) to promote germination. Repression of AtGLR3.5 impairs cytosolic Ca2+ concentration elevation, significantly delays germination, and enhances ABA sensitivity in seeds, whereas overexpression of AtGLR3.5 results in earlier germination and reduced seed sensitivity to ABA. Furthermore, we show that Ca2+ suppresses the expression of ABSCISIC ACID INSENSITIVE4 (ABI4), a key transcription factor involved in ABA response in seeds, and that ABI4 plays a fundamental role in modulation of Ca2+-dependent germination. Taken together, our results provide molecular genetic evidence that AtGLR3.5-mediated Ca2+ influx stimulates seed germination by antagonizing the inhibitory effects of ABA through suppression of ABI4. These findings establish, to our knowledge, a new and pivotal role of the plant glutamate receptor homolog and Ca2+ signaling in germination control and uncover the orchestrated modulation of the AtGLR3.5-mediated Ca2+ signal and ABA signaling via ABI4 to fine-tune the crucial developmental process, germination, in Arabidopsis.


Subject(s)
Abscisic Acid/metabolism , Arabidopsis Proteins/metabolism , Arabidopsis/physiology , Calcium/metabolism , Gene Expression Regulation, Plant , Plant Growth Regulators/metabolism , Receptors, Glutamate/metabolism , Arabidopsis/genetics , Arabidopsis Proteins/genetics , Cytosol/metabolism , Genes, Reporter , Germination , Models, Biological , Mutation , Receptors, Glutamate/genetics , Seeds/genetics , Seeds/physiology , Signal Transduction , Transcription Factors/genetics , Transcription Factors/metabolism
17.
Plant Cell Rep ; 34(2): 277-89, 2015 Feb.
Article in English | MEDLINE | ID: mdl-25410251

ABSTRACT

KEY MESSAGE: The Arabidopsis U-box E3 Ub ligase AtPUB30 participates in the salt stress tolerance as a negative factor in an ABA-independent manner during germination. Based on the in silico expression data, the U-box protein 30 (AtPUB30) from Arabidopsis thaliana was identified as a gene that responds to salt stress. The deduced AtPUB30 protein consists of 448 amino acids with a single U-box motif and five ARM-repeat domains. An in vitro self-ubiquitination assay demonstrated that bacterially expressed AtPUB30 exhibited E3 ubiquitin (Ub) ligase activity and that the U-box domain was essential for the activity. Real-time qRT-PCR and promoter-GUS analyses showed that AtPUB30 was induced by high salinity, but not by drought, cold, or abscisic acid (ABA), in roots but not in shoots. These results suggest that AtPUB30 is an Arabidopsis U-box E3 Ub ligase, the expression of which is selectively enhanced by salt stress in roots. T-DNA-inserted loss-of-function atpub30 mutant plants (atpub30-1 and atpub30-2) were more tolerant to salt stress in the germination stage, as identified by radicle emergence, cotyledon opening, and more vigorous early root growth relative to wild-type plants. Thus, it is likely that AtPUB30 plays a negative role in high salinity tolerance in the germination process. Wild type and mutant plants displayed very similar germination rates when treated with ABA, suggesting that the action of AtPUB30 in the germination stage is ABA independent. The post-germination growth of NaCl-stressed wild type and mutant plants were indistinguishable. Overall, our data suggest that the Arabidopsis U-box E3 Ub ligase AtPUB30 participates in the salt stress tolerance as a negative factor in the germination stage in root tissues.


Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis/physiology , Gene Expression Regulation, Plant , Sodium Chloride/pharmacology , Amino Acid Sequence , Arabidopsis/drug effects , Arabidopsis/genetics , Arabidopsis Proteins/metabolism , Gene Expression , Gene Knockout Techniques , Genes, Reporter , Germination , Molecular Sequence Data , Mutagenesis, Insertional , Phenotype , Phylogeny , Plant Roots/drug effects , Plant Roots/genetics , Plant Roots/physiology , Plants, Genetically Modified , Promoter Regions, Genetic/genetics , Salt Tolerance , Sequence Alignment , Stress, Physiological , Suppression, Genetic , Ubiquitin-Protein Ligases/genetics , Ubiquitin-Protein Ligases/metabolism
18.
Analyst ; 139(20): 5079-85, 2014 Oct 21.
Article in English | MEDLINE | ID: mdl-25109271

ABSTRACT

Advances in single cell analysis techniques have demonstrated cell-to-cell variability in both homogeneous and heterogeneous cell populations strengthening our understanding of multicellular organisms and individual cell behaviour. However, additional tools are needed for non-targeted metabolic analysis of live single cells in their native environment. Here, we combine capillary microsampling with electrospray ionization (ESI) mass spectrometry (MS) and ion mobility separation (IMS) for the analysis of various single A. thaliana epidermal cell types, including pavement and basal cells, and trichomes. To achieve microsampling of different cell types with distinct morphology, custom-tailored microcapillaries were used to extract the cell contents. To eliminate the isobaric interferences and enhance the ion coverage in single cell analysis, a rapid separation technique, IMS, was introduced that retained ions based on their collision cross sections. For each cell type, the extracted cell material was directly electrosprayed resulting in ∼200 peaks in ESI-MS and ∼400 different ions in ESI-IMS-MS, the latter representing a significantly enhanced coverage. Based on their accurate masses and tandem MS, 23 metabolites and lipids were tentatively identified. Our results indicated that profound metabolic differences existed between the trichome and the other two cell types but differences between pavement and basal cells were hard to discern. The spectra indicated that in all three A. thaliana cell types the phenylpropanoid metabolism pathway had high coverage. In addition, metabolites from the subpathway, sinapic acid ester biosynthesis, were more abundant in single pavement and basal cells, whereas compounds from the kaempferol glycoside biosynthesis pathway were present at significantly higher level in trichomes. Our results demonstrate that capillary microsampling coupled with ESI-IMS-MS captures metabolic differences between A. thaliana epidermal cell types, paving the way for the non-targeted analysis of single plant cells and subcellular compartments.


Subject(s)
Metabolomics/instrumentation , Metabolomics/methods , Plant Cells/chemistry , Spectrometry, Mass, Electrospray Ionization , Arabidopsis/chemistry , Arabidopsis/metabolism , Isomerism , Plant Cells/metabolism , Plant Leaves/chemistry , Plant Leaves/cytology , Plant Leaves/metabolism , Single-Cell Analysis
19.
Plant J ; 79(2): 322-33, 2014 Jul.
Article in English | MEDLINE | ID: mdl-24891222

ABSTRACT

Faced with declining soil-water potential, plants synthesize abscisic acid (ABA), which then triggers stomatal closure to conserve tissue moisture. Closed stomates, however, also create several physiological dilemmas. Among these, the large CO2 influx required for net photosynthesis will be disrupted. Depleting CO2 in the plant will in turn bias stomatal opening by suppressing ABA sensitivity, which then aggravates transpiration further. We have investigated the molecular basis of how C3 plants resolve this H2 O-CO2 conflicting priority created by stomatal closure. Here, we have identified in Arabidopsis thaliana an early drought-induced spermidine spermine-N(1) -acetyltransferase homolog, which can slow ABA-mediated stomatal closure. Evidence from genetic, biochemical and physiological analyses has revealed that this protein does so by acetylating the metabolite 1,3-diaminopropane (DAP), thereby turning on the latter's intrinsic activity. Acetylated DAP triggers plasma membrane electrical and ion transport properties in an opposite way to those by ABA. Thus in adapting to low soil-water availability, acetyl-DAP could refrain stomates from complete closure to sustain CO2 diffusion to photosynthetic tissues.


Subject(s)
Abscisic Acid/metabolism , Arabidopsis/metabolism , Diamines/metabolism , Droughts , Plant Stomata/metabolism , Arabidopsis Proteins/metabolism , Gene Expression Regulation, Plant , Signal Transduction
20.
Nucleic Acids Res ; 42(Web Server issue): W198-204, 2014 Jul.
Article in English | MEDLINE | ID: mdl-24878919

ABSTRACT

Pairwise comparison of data vectors represents a large part of computational biology, especially with the continuous increase in genome-wide approaches yielding more information from more biological samples simultaneously. Gene clustering for function prediction as well as analyses of signalling pathways and the time-dependent dynamics of a system are common biological approaches that often rely on large dataset comparison. Different metrics can be used to evaluate the similarity between entities to be compared, such as correlation coefficients and distances. While the latter offers a more flexible way of measuring potential biological relationships between datasets, the significance of any given distance is highly dependent on the dataset and cannot be easily determined. Monte Carlo methods are robust approaches for evaluating the significance of distance values by multiple random permutations of the dataset followed by distance calculation. We have developed R. S. WebTool (http://rswebtool.kwaklab.org), a user-friendly online server for random sampling-based evaluation of distance significances that features an array of visualization and analysis tools to help non-bioinformaticist users extract significant relationships from random noise in distance-based dataset analyses.


Subject(s)
Computational Biology/methods , Software , Algorithms , Gene Expression Profiling , Internet , Monte Carlo Method
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