Wild-type Columbia Research Articles

Molecular constraints on tolerance-resistance trade-offs: Is there a cost?

Plants possess myriad defenses against their herbivores, including constitutive and inducible chemical compounds and regrowth strategies known as tolerance. Recent studies have shown that plant tolerance and resistance are positively associated given they are co-localized in the same molecular pathway, the oxidative pentose phosphate pathway. However, given that both defensive strategies utilize carbon skeletons from a shared resource pool in the oxidative pentose phosphate pathway there are likely costs in maintaining both resistance-tolerance strategies. Here we investigate fitness costs in maintaining both strategies by utilizing a double knockout of cyp79B2 and cyp79B3, key enzymes in the biosynthetic process of indole glucosinolates, which convert tryptophan to indole-3-acetaldoxime (IAOx) and is further used to produce indole glucosinolates. These mutant plants are devoid of any indole glucosinolates thus reducing plant resistance. Results show that knocking out indole glucosinolate production and thus one of the resistance pathways leads to an approximate 94% increase in fitness compensation shifting the undercompensating wild-type Columbia-0 to an overcompensating genotype following damage. We discuss the potential mechanistic basis for the observed patterns.

Arabidopsis plasma membrane intrinsic protein (AtPIP2;1) is implicated in a salinity conditional influence on seed germination.

Dynamic changes in aquaporin gene expression occur during seed germination. One example is the ~30-fold increase in Arabidopsis thaliana PIP2;1 transcripts within 24h of seed imbibition. To investigate whether AtPIP2;1 can influence seed germination wild-type Columbia-0, single (Atpip2;1 ) and double (Atpip2;1-Atpip2;2 ) loss-of-function mutants, along with transgenic 2x35S::AtPIP2;1 over-expressing (OE) lines and null-segregant controls, were examined. The various genotypes were germinated in control and saline (75mM NaCl treatment) conditions and tested for germination efficiency, imbibed seed maximum cross sectional (MCS) area, imbibed seed mass, and seed Na+ and K+ content. Seed lacking functional AtPIP2;1 and/or AtPIP2;2 proteins or constitutively over-expressing AtPIP2;1 , had delayed germination in saline conditions relative to wild-type and null-segregant seed, respectively. Exposure to saline germination conditions resulted in Atpip2;1 mutants having greater imbibed seed mass and less accumulated Na+ than wild-type, whereas lines over-expressing AtPIP2;1 had reduced imbibed seed mass and greater seed K+ content than null-segregant control seed. The results imply a role for AtPIP2;1 in seed germination processes, whether directly through its capacity for water and ion transport or H2 O2 signalling, or indirectly through potentially triggering dynamic differential regulation of other aquaporins expressed during germination. Future research will aid in dissecting the aquaporin functions influencing germination and may lead to novel solutions for optimising germination in sub-optimal conditions, such as saline soils.

Nondestructive circadian profiling of starch content in fresh intact Arabidopsis leaf with two-photon fluorescence and second-harmonic generation imaging

Plant chloroplasts conduct photosynthesis to convert solar energy into sugars for the carbon source essential for cell living and growth during the day. One fraction of photosynthetic products is stored in chloroplasts by forming starch granules to continue the provision of carbon energy during the night. Currently, profiling the starch temporal pattern requires either: (i) sacrificing the leaves, or (ii) generating transgenic plants at the risk of changing the metabolisms by incorporating a genetically modified granule-bound starch synthase (GBSS). In this paper, we demonstrated a nondestructive method using two-photon fluorescence (TPF) and second-harmonic generation (SHG) imaging to quantify starch granules within chloroplasts of fresh intact leaves across a day-night cycle. We did so using two Arabidopsis lines having normal and excess starch contents: wild-type (Columbia-0) and starch excess 1 (sex1). The starch granules were visualized by SHG imaging, while the chloroplasts in mesophyll cells were visualized by TPF imaging. Our results provided micron scale spatial resolution of starch distribution within leaves and showed starch circadian patterns consistent with those profiled by enzymatic assays in previous studies. We demonstrated that TPF-SHG imaging is a potential tool for revealing the real-time heterogeneity of starch circadian rhythm in leaf cells, without the need for destructive sample preparation.

Investigating root architectural differences in lines of Arabidopsis thaliana. L. with altered stomatal density using high resolution X-Ray synchrotron imaging

PurposeFreshwater is an increasingly scarce natural resource, essential for agricultural production. As plants consume 70% of the world’s freshwater, a reduction in their water use would greatly reduce global water scarcity. Plants with improved Water Use Efficiency (WUE) such as those with altered expression of the Epidermal Patterning Factor (EPF) family of genes regulating stomatal density, could help reduce plant water footprint. Little however, is known about how this modification in Arabidopsis thaliana. L. affects root architectural development in soil, thus we aim to improve our understanding of root growth when stomatal density is altered.MethodsWe used X-Ray synchrotron and neutron imaging to measure in three dimensions, the root system architecture (RSA) of Arabidopsis thaliana. L. plants of three different genotypes, namely that of the wild type Columbia (Col 0) and two different EPF mutants, EPF2OE and epf2-1 (which show reduced and increased stomatal density, respectively). We also used the total biomass and carbon isotope discrimination (Δ) methods to determine how WUE varies in these genotypes when grown in a sandy loam soil under controlled conditions.ResultsOur results confirm that the EPF2OE line had superior WUE as compared to the wild type using both the Δ and total biomass method. The epf2-1 mutant, on the other hand, had significantly reduced WUE using the Δ but not with the biomass method. In terms of root growth, the RSAs of the different genotypes had no significant difference between each other. There was also no significant difference in rhizosphere porosity around their roots as compared to bulk soil for all genotypes.ConclusionOur results indicate that the EPF mutation altering stomatal density in Arabidopsis thaliana. L. plants did not have an adverse effect on root characteristics thus their wide adoption to reduce the global freshwater footprint is unlikely to compromise their soil foraging ability.

Lipidomic Analysis of Arabidopsis T-DNA Insertion Lines Leads to Identification and Characterization of C-Terminal Alterations in FATTY ACID DESATURASE 6.

Mass-spectrometry-based screening of lipid extracts of wounded and unwounded leaves from a collection of 364 Arabidopsis thaliana T-DNA insertion lines produced lipid profiles that were scored on the number and significance of their differences from the leaf lipid profiles of wild-type plants. The analysis identified Salk_109175C, which displayed alterations in leaf chloroplast glycerolipid composition, including a decreased ratio between two monogalactosyldiacylglycerol (MGDG) molecular species, MGDG(18:3/16:3) and MGDG(18:3/18:3). Salk_109175C has a confirmed insertion in the At5g64790 locus; the insertion did not co-segregate with the recessive lipid phenotype in the F2 generation of a wild-type (Columbia-0) × Salk_109175C cross. The altered lipid compositional phenotype mapped to the At4g30950 locus, which encodes the plastidial ω-6 desaturase FATTY ACID DESATURASE 6 (FAD6). Sequencing revealed a splice-site mutation, leading to the in-frame deletion of 13 amino acids near the C-terminal end of the 448 amino acid protein. Heterologous expression in yeast showed that this deletion eliminates desaturase activity and reduces protein stability. Sequence comparison across species revealed that several amino acids within the deletion are conserved in plants and cyanobacteria. Individual point mutations in four conserved residues resulted in 77–97% reductions in desaturase activity, while a construct with all four alanine substitutions lacked activity. The data suggest that the deleted region of FAD6, which is on the C-terminal side of the four putative transmembrane segments and the histidine boxes putatively involved in catalysis, is critical for FAD6 function.

Suppression of Cortical Microtubule Reorientation and Stimulation of Cell Elongation in Arabidopsis Hypocotyls under Microgravity Conditions in Space.

How microgravity in space influences plant cell growth is an important issue for plant cell biology as well as space biology. We investigated the role of cortical microtubules in the stimulation of elongation growth in Arabidopsis (Arabidopsis thaliana) hypocotyls under microgravity conditions with the Resist Tubule space experiment. The epidermal cells in the lower half of the hypocotyls of wild-type Columbia were longer in microgravity than at on-orbit 1 g, which precipitated an increase in the entire hypocotyl length. In the apical region, cortical microtubules adjacent to the outer tangential wall were predominantly transverse to the long axis of the cell, whereas longitudinal microtubules were predominant in the basal region. In the 9th to 12th epidermal cells (1 to 3 mm) from the tip, where the modification of microtubule orientation from transverse to longitudinal directions (reorientation) occurred, cells with transverse microtubules increased, whereas those with longitudinal microtubules decreased in microgravity, and the average angle with respect to the transverse cell axis decreased, indicating that the reorientation was suppressed in microgravity. The expression of tubulin genes was suppressed in microgravity. These results suggest that under microgravity conditions, the expression of genes related to microtubule formation was downregulated, which may cause the suppression of microtubule reorientation from transverse to longitudinal directions, thereby stimulating cell elongation in Arabidopsis hypocotyls.

Exogenous sucrose influences KEA1 and KEA2 to regulate abscisic acid-mediated primary root growth in Arabidopsis

Arabidopsis K+-efflux antiporter (KEA)1 and KEA2 are chloroplast inner envelope membrane K+/H+ antiporters that play an important role in plastid development and seedling growth. However, the function of KEA1 and KEA2 during early seedling development is poorly understood. In this work, we found that in Arabidopsis, KEA1 and KEA2 mediated primary root growth by regulating photosynthesis and the ABA signaling pathway. Phenotypic analyses revealed that in the absence of sucrose, the primary root length of the kea1kea2 mutant was significantly shorter than that of the wild-type Columbia-0 (Col-0) plant. However, this phenotype could be remedied by the external application of sucrose. Meanwhile, HPLC–MS/MS results showed that in sucrose-free medium, ABA accumulation in the kea1kea2 mutant was considerably lower than that in Col-0. Transcriptome analysis revealed that many key genes involved in ABA signals were repressed in the kea1kea2 mutant. We concluded that KEA1 and KEA2 deficiency not only affected photosynthesis but was also involved in primary root growth likely through an ABA-dependent manner. This study confirmed the new function of KEA1 and KEA2 in affecting primary root growth.

Identification of Elements Responsible for Maternally-Silenced Imprinted Gene Expression of Upward Curly Leaf1, an F-box Protein Gene that Regulates Curly Leaf in Arabidopsis

Upward Curly Leaf 1 (UCL1) is an Arabidopsis thaliana E3 ligase that targets the Curly Leaf (CLF) SET-domain polycomb-group (PcG) protein for degradation via the ubiquitin-26S proteasome system. UCL1 is a paternally-expressed imprinted gene in the endosperm. To precisely locate the promoter elements required for UCL1 imprinting pattern, various gene constructs were created in which the imprinting control region (ICR), endosperm-specific expression (ENSE) element, and/or the linker sequence were altered. By fusing these constructs with a GUS reporter gene, GUS expression patterns were monitored after reciprocal crosses with wild-type Columbia-0 allowing the determination of parent-of-origin expression. Analysis of publicly-available data on the UCL1 promoter region facilitated the search for allele-specific DNA and H3K27 methylation patterns. Overall, three promoter elements are required for maternal repression of UCL1; the ICR sequence located from − 2.5 to − 2.4 kb upstream of the translation start site, a differentially methylated region 2 (DMR2) that overlaps the short ATLINE1-1 transposable element in the linker region, and a minimal 271 bp ENSE element. In addition, DNA methylation patterns in the DMR2 contribute to the repression of the maternal UCL1 allele. Our findings would help to understand how parent-of-origin epigenetic patterns are created and maintained in the endosperm.

CIRCADIAN CLOCK-ASSOCIATED1 Controls Resistance to Aphids by Altering Indole Glucosinolate Production.

CIRCADIAN CLOCK-ASSOCIATED1 (CCA1), a well-known central circadian clock regulator, coordinates plant responses to environmental challenges. Its daily rhythmic expression in Arabidopsis (Arabidopsis thaliana) confers host resistance to the caterpillar Trichoplusia ni However, it is unclear whether CCA1 plays a role in defense against phloem sap-feeding aphids. In this study, we showed that green peach aphid (Myzus persicae) displayed an intrinsic circadian feeding rhythm. Under constant light, wild-type Columbia-0 (Col-0) Arabidopsis plants coentrained with aphids in the same light/dark cycles exhibited greater antixenotic activity than plants preentrained in the opposite cycle from the aphids. Consistently, circadian mutants cca1-1, cca1-11, lhy-21, ztl-1, ztl-4, and lux-2 suffered more severe damage than Col-0 plants when infested by aphids, suggesting that the Arabidopsis circadian clock plays a defensive role. However, the arrhythmic CCA1 overexpression line (CCA1-OX) displayed strong antixenotic and antibiotic activities despite its loss of circadian regulation. Aphids feeding on CCA1-OX plants exhibited lower reproduction and smaller body size and weight than those on Col-0. Apparently, CCA1 regulates both clock-dependent and -independent defense responses. Systematic investigation based on bioinformatics analyses indicated that resistance to aphids in CCA1-OX plants was due primarily to heightened basal indole glucosinolate levels. Interestingly, aphid feeding induced alternatively spliced intron-retaining CCA1a/b transcripts, which are normally expressed at low levels, whereas expression of the major fully spliced CCA1 transcript remained largely unchanged. We hypothesize that posttranscriptional modulation of CCA1 expression upon aphid infestation maximizes the potential of circadian-mediated defense and stress tolerance while ensuring normal plant development.

The Photoperiodic Flowering Time Regulator FKF1 Negatively Regulates Cellulose Biosynthesis.

Cellulose synthesis is precisely regulated by internal and external cues, and emerging evidence suggests that light regulates cellulose biosynthesis through specific light receptors. Recently, the blue light receptor CRYPTOCHROME 1 (CRY1) was shown to positively regulate secondary cell wall biosynthesis in Arabidopsis (Arabidopsis thaliana). Here, we characterize the role of FLAVIN-BINDING KELCH REPEAT, F-BOX 1 (FKF1), another blue light receptor and well-known photoperiodic flowering time regulator, in cellulose biosynthesis. A phenotype suppression screen using a cellulose deficient mutant cesa1aegeus,cesa3ixr1-2 (c1,c3), which carries nonlethal point mutations in CELLULOSE SYNTHASE A 1 (CESA1) and CESA3, resulted in identification of the phenotype-restoring large leaf (llf) mutant. Next-generation mapping using the whole genome resequencing method identified the llf locus as FKF1 FKF1 was confirmed as the causal gene through observation of the llf phenotype in an independent triple mutant c1,c3,fkf1-t carrying a FKF1 T-DNA insertion mutant. Moreover, overexpression of FKF1 in llf plants restored the c1,c3 phenotype. The fkf1 mutants showed significant increases in cellulose content and CESA gene expression compared with that in wild-type Columbia-0 plants, suggesting a negative role of FKF1 in cellulose biosynthesis. Using genetic, molecular, and phenocopy and biochemical evidence, we have firmly established the role of FKF1 in regulation of cellulose biosynthesis. In addition, CESA expression analysis showed that diurnal expression patterns of CESAs are FKF1 independent, whereas their circadian expression patterns are FKF1 dependent. Overall, our work establishes a role of FKF1 in the regulation of cell wall biosynthesis in Arabidopsis.

The Distinct Roles of Class I and II RPD3-Like Histone Deacetylases in Salinity Stress Response.

Histone acetylation is an essential process in the epigenetic regulation of diverse biological processes, including environmental stress responses in plants. Previously, our research group identified a histone deacetylase (HDAC) inhibitor (HDI) that confers salt tolerance in Arabidopsis (Arabidopsis thaliana). In this study, we demonstrate that class I HDAC (HDA19) and class II HDACs (HDA5/14/15/18) control responses to salt stress through different pathways. The screening of 12 different selective HDIs indicated that seven newly reported HDIs enhance salt tolerance. Genetic analysis, based on a pharmacological study, identified which HDACs function in salinity stress tolerance. In the wild-type Columbia-0 background, hda19 plants exhibit tolerance to high-salinity stress, while hda5/14/15/18 plants exhibit hypersensitivity to salt stress. Transcriptome analysis revealed that the effect of HDA19 deficiency on the response to salinity stress is distinct from that of HDA5/14/15/18 deficiencies. In hda19 plants, the expression levels of stress tolerance-related genes, late embryogenesis abundant proteins that prevent protein aggregation and positive regulators such as ABI5 and NAC019 in abscisic acid signaling, were induced strongly relative to the wild type. Neither of these elements was up-regulated in the hda5/14/15/18 plants. The mutagenesis of HDA19 by genome editing in the hda5/14/15/18 plants enhanced salt tolerance, suggesting that suppression of HDA19 masks the phenotype caused by the suppression of class II HDACs in the salinity stress response. Collectively, our results demonstrate that HDIs that inhibit class I HDACs allow the rescue of plants from salinity stress regardless of their selectivity, and they provide insight into the hierarchal regulation of environmental stress responses through HDAC isoforms.

Growth in spaceflight hardware results in alterations to the transcriptome and proteome

The Biological Research in Canisters (BRIC) hardware has been used to house many biology experiments on both the Space Transport System (STS, commonly known as the space shuttle) and the International Space Station (ISS). However, microscopic examination of Arabidopsis seedlings by Johnson et al. (2015) indicated the hardware itself may affect cell morphology. The experiment herein was designed to assess the effects of the BRIC-Petri Dish Fixation Units (BRIC-PDFU) hardware on the transcriptome and proteome of Arabidopsis seedlings. To our knowledge, this is the first transcriptomic and proteomic comparison of Arabidopsis seedlings grown with and without hardware. Arabidopsis thaliana wild-type Columbia (Col-0) seeds were sterilized and bulk plated on forty-four 60 mm Petri plates, of which 22 were integrated into the BRIC-PDFU hardware and 22 were maintained in closed containers at Ohio University. Seedlings were grown for approximately 3 days, fixed with RNAlater® and stored at -80 °C prior to RNA and protein extraction, with proteins separated into membrane and soluble fractions prior to analysis. The RNAseq analysis identified 1651 differentially expressed genes; MS/MS analysis identified 598 soluble and 589 membrane proteins differentially abundant both at p < .05. Fold enrichment analysis of gene ontology terms related to differentially expressed transcripts and proteins highlighted a variety of stress responses. Some of these genes and proteins have been previously identified in spaceflight experiments, indicating that these genes and proteins may be perturbed by both conditions.

Arabidopsis thaliana miRNAs promote embryo pattern formation beginning in the zygote

miRNAs are essential regulators of cell identity, yet their role in early embryo development in plants remains largely unexplored. To determine the earliest stage at which miRNAs act to promote pattern formation in embryogenesis, we examined a series of mutant alleles in the Arabidopsis thaliana miRNA biogenesis enzymes DICER-LIKE 1 (DCL1), SERRATE (SE), and HYPONASTIC LEAVES 1 (HYL1). Cellular and patterning defects were observed in dcl1, se and hyl1 embryos from the zygote through the globular stage of embryogenesis. To identify miRNAs that are expressed in early embryogenesis, we sequenced mRNAs from globular stage Columbia wild type (wt) and se-1 embryos, and identified transcripts potentially corresponding to 100 miRNA precursors. Considering genome location and transcript increase between wt and se-1, 39 of these MIRNAs are predicted to be bona fide early embryo miRNAs. Among these are conserved miRNAs such as miR156, miR159, miR160, miR161, miR164, miR165, miR166, miR167, miR168, miR171, miR319, miR390 and miR394, as well as miRNAs whose function has never been characterized. Our analysis demonstrates that miRNAs promote pattern formation beginning in the zygote, and provides a comprehensive dataset for functional studies of individual miRNAs in Arabidopsis embryogenesis.

Effect of nitric oxide donor on salt resistance of Arabidopsis jin1 mutants and wild-type plants

The effect of NO donor sodium nitroprusside (SNP) on salt resistance of 4-week-old Arabidopsis thaliana L. wild-type Columbia-0 (Col-0) plants and jin1 mutants defective in the jasmonate signaling have been investigated. As affected by 0.5 mM, SNP salt resistance of wild-type plants rose, which was exhibited in a smaller growth inhibition and preserving the pool of photosynthetic pigments after salt stress (200 mM NaCl). The positive effect of SNP leveled by treatment of plant with NO scavenger: 0.5 mM PTIO (2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide). However, SNP treatment had no significant effect on the salt tolerance of jin1 genotype plants. In the case of wild-type plants but not jin1 mutants, treatment by NO donor increased activity of antioxidant enzymes—superoxide dismutase, guaiacol peroxidase, and catalase—in the leaves, which was especially noticeable in salt stress conditions. In wild-type plants treated by NO donor, proline content in the leaves after salt stress was lower and sugar content was higher than that in the untreated ones. In jin1 mutants, NO donor treatment resulted in a significant increase in proline content in the leaves under salt stress, without changing sugar content. A conclusion was made on the participation of transcript factor JIN1/MYC2 in NO-dependent induction of some plant defense responses to salt stress.

FUM2, a Cytosolic Fumarase, Is Essential for Acclimation to Low Temperature in Arabidopsis thaliana

Although cold acclimation is a key process in plants from temperate climates, the mechanisms sensing low temperature remain obscure. Here, we show that the accumulation of the organic acid fumaric acid, mediated by the cytosolic fumarase FUM2, is essential for cold acclimation of metabolism in the cold-tolerant model species Arabidopsis (Arabidopsis thaliana). A nontargeted metabolomic approach, using gas chromatography-mass spectrometry, identifies fumarate as a key component of the cold response in this species. Plants of T-DNA insertion mutants, lacking FUM2, show marked differences in their response to cold, with contrasting responses both in terms of metabolite concentrations and gene expression. The fum2 plants accumulated higher concentrations of phosphorylated sugar intermediates and of starch and malate. Transcripts for proteins involved in photosynthesis were markedly down-regulated in fum2.2 but not in wild-type Columbia-0. Plants of fum2 show a complete loss of the ability to acclimate photosynthesis to low temperature. We conclude that fumarate accumulation plays an essential role in low temperature sensing in Arabidopsis, either indirectly modulating metabolic or redox signals or possibly being itself directly involved in cold sensing.

Arabidopsis NATA1 Acetylates Putrescine and Decreases Defense-Related Hydrogen Peroxide Accumulation.

Biosynthesis of the polyamines putrescine, spermidine, and spermine is induced in response to pathogen infection of plants. Putrescine, which is produced from Arg, serves as a metabolic precursor for longer polyamines, including spermidine and spermine. Polyamine acetylation, which has important regulatory functions in mammalian cells, has been observed in several plant species. Here we show that Arabidopsis (Arabidopsis thaliana) N-ACETYLTRANSFERASE ACTIVITY1 (NATA1) catalyzes acetylation of putrescine to N-acetylputrescine and thereby competes with spermidine synthase for a common substrate. NATA1 expression is strongly induced by the plant defense signaling molecule jasmonic acid and coronatine, an effector molecule produced by DC3000, a Pseudomonas syringae strain that initiates a virulent infection in Arabidopsis ecotype Columbia-0. DC3000 growth is reduced in nata1 mutant Arabidopsis, suggesting a role for NATA1-mediated putrescine acetylation in suppressing antimicrobial defenses. During infection by P. syringae and other plant pathogens, polyamine oxidases use spermidine and spermine as substrates for the production of defense-related H2O2 Compared to wild-type Columbia-0 Arabidopsis, the response of nata1mutants to P. syringae infection includes reduced accumulation of acetylputrescine, greater abundance of nonacetylated polyamines, elevated H2O2 production by polyamine oxidases, and higher expression of genes related to pathogen defense. Together, these results are consistent with a model whereby P. syringae growth is improved in a targeted manner through coronatine-induced putrescine acetylation by NATA1.

Content of Osmolytes and Flavonoids under Salt Stress in Arabidopsis thaliana Plants Defective in Jasmonate Signaling

The effects of the salt stress (200 mM NaCl) and exogenous jasmonic acid (JA) on levels of osmolytes and flavonoids in leaves of four-week-old Arabidopsis thaliana L. plants of the wild-type (WT) Columbia-0 (Col-0) and the mutant jin1 (jasmonate insensitive 1) with impaired jasmonate signaling were studied. The increase in proline content caused by the salt stress was higher in the Col-0 plants than in the mutant jin1. This difference was especially marked if the plants had been pretreated with exogenous 0.1 µM JA. The sugar content increased in response to the salt stress in the JA-treated WT plants but decreased in the jin1 mutant. Leaf treatment with JA of the WT plants but not mutant defective in jasmonate signaling also enhanced the levels of anthocyanins and flavonoids absorbed in UV-B range. The presence of JA increased salinity resistance of the Col-0 plants, since the accumulation of lipid peroxidation products and growth inhibition caused by NaCl were less pronounced. Under salt stress, JA almost did not render a positive effect on the jin1 plants. It is concluded that the protein JIN1/MYC2 is involved in control of protective systems under salt stress.

A Predictive Coexpression Network Identifies Novel Genes Controlling the Seed-to-Seedling Phase Transition in Arabidopsis thaliana.

The transition from a quiescent dry seed to an actively growing photoautotrophic seedling is a complex and crucial trait for plant propagation. This study provides a detailed description of global gene expression in seven successive developmental stages of seedling establishment in Arabidopsis (Arabidopsis thaliana). Using the transcriptome signature from these developmental stages, we obtained a coexpression gene network that highlights interactions between known regulators of the seed-to-seedling transition and predicts the functions of uncharacterized genes in seedling establishment. The coexpressed gene data sets together with the transcriptional module indicate biological functions related to seedling establishment. Characterization of the homeodomain leucine zipper I transcription factor AtHB13, which is expressed during the seed-to-seedling transition, demonstrated that this gene regulates some of the network nodes and affects late seedling establishment. Knockout mutants for athb13 showed increased primary root length as compared with wild-type (Columbia-0) seedlings, suggesting that this transcription factor is a negative regulator of early root growth, possibly repressing cell division and/or cell elongation or the length of time that cells elongate. The signal transduction pathways present during the early phases of the seed-to-seedling transition anticipate the control of important events for a vigorous seedling, such as root growth. This study demonstrates that a gene coexpression network together with transcriptional modules can provide insights that are not derived from comparative transcript profiling alone.

Nitrate reductase mutation alters potassium nutrition as well as nitric oxide-mediated control of guard cell ion channels in Arabidopsis.

Maintaining potassium (K(+) ) nutrition and a robust guard cell K(+) inward channel activity is considered critical for plants' adaptation to fluctuating and challenging growth environment. ABA induces stomatal closure through hydrogen peroxide and nitric oxide (NO) along with subsequent ion channel-mediated loss of K(+) and anions. However, the interactions of NO synthesis and signalling with K(+) nutrition and guard cell K(+) channel activities have not been fully explored in Arabidopsis. Physiological and molecular techniques were employed to dissect the interaction of nitrogen and potassium nutrition in regulating stomatal opening, CO2 assimilation and ion channel activity. These data, gene expression and ABA signalling transduction were compared in wild-type Columbia-0 (Col-0) and the nitrate reductase mutant nia1nia2. Growth and K(+) nutrition were impaired along with stomatal behaviour, membrane transport, and expression of genes associated with ABA signalling in the nia1nia2 mutant. ABA-inhibited K(+) in current and ABA-enhanced slow anion current were absent in nia1nia2. Exogenous NO restored regulation of these channels for complete stomatal closure in nia1nia2. While NO is an important signalling component in ABA-induced stomatal closure in Arabidopsis, our findings demonstrate a more complex interaction associating potassium nutrition and nitrogen metabolism in the nia1nia2 mutant that affects stomatal function.

PhenoVein-A Tool for Leaf Vein Segmentation and Analysis.

Precise measurements of leaf vein traits are an important aspect of plant phenotyping for ecological and genetic research. Here, we present a powerful and user-friendly image analysis tool named phenoVein. It is dedicated to automated segmenting and analyzing of leaf veins in images acquired with different imaging modalities (microscope, macrophotography, etc.), including options for comfortable manual correction. Advanced image filtering emphasizes veins from the background and compensates for local brightness inhomogeneities. The most important traits being calculated are total vein length, vein density, piecewise vein lengths and widths, areole area, and skeleton graph statistics, like the number of branching or ending points. For the determination of vein widths, a model-based vein edge estimation approach has been implemented. Validation was performed for the measurement of vein length, vein width, and vein density of Arabidopsis (Arabidopsis thaliana), proving the reliability of phenoVein. We demonstrate the power of phenoVein on a set of previously described vein structure mutants of Arabidopsis (hemivenata, ondulata3, and asymmetric leaves2-101) compared with wild-type accessions Columbia-0 and Landsberg erecta-0. phenoVein is freely available as open-source software.