ABSTRACT
Foliar pigmentation patterns vary among plant species and growth conditions. In this study, we utilize hyperspectral imaging to assess foliar pigmentation in the bryophyte Marchantia polymorpha under nutrient stress and identify associated genetic factors. Using singular value decomposition (SVD) for feature selection, we quantitate color variations induced by deficiencies in phosphate, nitrate, magnesium, calcium, and iron. Pseudo-colored thallus images show that disrupting MpWRKY10 causes irregular pigmentation with auronidin accumulation. Transcriptomic profiling shows that MpWRKY10 regulates phenylpropanoid pathway enzymes and R2R3-MYB transcription factors during phosphate deficiency, with MpMYB14 upregulation preceding pigment accumulation. MpWRKY10 is downregulated in older, pigmented thalli under phosphate deficiency but maintained in young thalli, where it suppresses pigmentation genes. This downregulation is absent in pigmented thalli due to aging. Comparative transcriptome analysis suggests similar WRKY and MYB roles in nutrient response and pigmentation in red-leaf lettuce, alluding to conserved genetic factors controlling foliar pigmentation patterns under nutrient deficiency.
Subject(s)
Gene Expression Regulation, Plant , Hyperspectral Imaging , Marchantia , Pigmentation , Plant Proteins , Pigmentation/genetics , Marchantia/genetics , Marchantia/metabolism , Plant Proteins/metabolism , Plant Proteins/genetics , Hyperspectral Imaging/methods , Plant Leaves/metabolism , Plant Leaves/genetics , Transcription Factors/metabolism , Transcription Factors/geneticsABSTRACT
mRNA translation relies on identifying translation initiation sites (TISs) in mRNAs. Alternative TISs are prevalent across plant transcriptomes, but the mechanisms for their recognition are unclear. Using ribosome profiling and machine learning, we developed models for predicting alternative TISs in the tomato (Solanum lycopersicum). Distinct feature sets were predictive of AUG and nonAUG TISs in 5' untranslated regions and coding sequences, including a novel CU-rich sequence that promoted plant TIS activity, a translational enhancer found across dicots and monocots, and humans and viruses. Our results elucidate the mechanistic and evolutionary basis of TIS recognition, whereby cis-regulatory RNA signatures affect start site selection. The TIS prediction model provides global estimates of TISs to discover neglected protein-coding genes across plant genomes. The prevalence of cis-regulatory signatures across plant species, humans, and viruses suggests their broad and critical roles in reprogramming the translational landscape.
Subject(s)
Eukaryota , Peptide Chain Initiation, Translational , Humans , Peptide Chain Initiation, Translational/genetics , Eukaryota/genetics , Plants/genetics , 5' Untranslated Regions , RNA, Messenger/genetics , Codon, InitiatorABSTRACT
The microbe-associated molecular pattern flg22 is recognized in a flagellin-sensitive 2-dependent manner in root tip cells. Here, we show a rapid and massive change in protein abundance and phosphorylation state of the Arabidopsis root cell proteome in WT and a mutant deficient in heterotrimeric G-protein-coupled signaling. flg22-induced changes fall on proteins comprising a subset of this proteome, the heterotrimeric G protein interactome, and on highly-populated hubs of the immunity network. Approximately 95% of the phosphorylation changes in the heterotrimeric G-protein interactome depend, at least partially, on a functional G protein complex. One member of this interactome is ATBα, a substrate-recognition subunit of a protein phosphatase 2A complex and an interactor to Arabidopsis thaliana Regulator of G Signaling 1 protein (AtRGS1), a flg22-phosphorylated, 7-transmembrane spanning modulator of the nucleotide-binding state of the core G-protein complex. A null mutation of ATBα strongly increases basal endocytosis of AtRGS1. AtRGS1 steady-state protein level is lower in the atbα mutant in a proteasome-dependent manner. We propose that phosphorylation-dependent endocytosis of AtRGS1 is part of the mechanism to degrade AtRGS1, thus sustaining activation of the heterotrimeric G protein complex required for the regulation of system dynamics in innate immunity. The PP2A(ATBα) complex is a critical regulator of this signaling pathway.
Subject(s)
Arabidopsis Proteins , Arabidopsis , Heterotrimeric GTP-Binding Proteins , RGS Proteins , Arabidopsis/metabolism , Phosphorylation , Arabidopsis Proteins/metabolism , Proteome/metabolism , RGS Proteins/chemistry , RGS Proteins/genetics , RGS Proteins/metabolism , Signal Transduction , Heterotrimeric GTP-Binding Proteins/metabolism , Flagellin/pharmacology , Flagellin/metabolism , Phosphoric Monoester Hydrolases/metabolismABSTRACT
Traditional methods for assessing plant health often lack the necessary attributes for continuous and non-destructive monitoring. In this pilot study, we present a novel technique utilizing a customized fiber optic probe based on attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR) with a contact force control unit for non-invasive and continuous plant health monitoring. We also developed a normalized difference mid-infrared reflectance index through statistical analysis of spectral features, enabling differentiation of drought and age conditions in plants. Our research aims to characterize phytochemicals and plant endogenous status optically, addressing the need for improved analytical measurement methods for in situ plant health assessment. The probe configuration was optimized with a triple-loop tip and a 3 N contact force, allowing sensitive measurements while minimizing leaf damage. By combining polycrystalline and chalcogenide fiber probes, a comprehensive wavenumber range analysis (4000-900 cm-1) was achieved. Results revealed significant variations in phytochemical composition among plant species, for example, red spinach with the highest polyphenolic content and green kale with the highest lignin content. Petioles displayed higher lignin and cellulose absorbance values compared to veins. The technique effectively monitored drought stress on potted green bok choy plants in situ, facilitating the quantification of changes in water content, antioxidant activity, lignin, and cellulose levels. This research represents the first demonstration of the potential of fiber optic ATR-FTIR probes for non-invasive and rapid plant health measurements, providing insights into plant health and advancements in quantitative monitoring for indoor farming practices, bioanalytical chemistry, and environmental sciences.
Subject(s)
Brassica , Lignin , Pilot Projects , Cellulose , Spectroscopy, Fourier Transform Infrared/methodsABSTRACT
Advanced precision agriculture requires the objective measurement of the structural and functional properties of plants. Biochemical profiles in leaves can differ depending on plant growing conditions. By quantitatively detecting these changes, farm production processes can be optimized to achieve high-yield, high-quality, and nutrient dense agricultural products. To enable the rapid and non-destructive detection on site, this study demonstrates the development of a new custom-designed portable handheld Vis-NIR spectrometer that collects leaf reflectance spectra, wirelessly transfers the spectral data through Bluetooth, and provides both raw spectral data and processed information. The spectrometer has two preprogramed methods: anthocyanin and chlorophyll quantification. Anthocyanin content of red and green lettuce estimated with the new spectrometer showed an excellent correlation coefficient of 0.84 with those determined by a destructive gold standard biochemical method. The differences in chlorophyll content were measured using leaf senescence as a case study. Chlorophyll Index calculated with the handheld spectrometer gradually decreased with leaf age as chlorophyll degrades during the process of senescence. The estimated chlorophyll values were highly correlated with those obtained from a commercial fluorescence-based chlorophyll meter with a correlation coefficient of 0.77. The developed portable handheld Vis-NIR spectrometer could be a simple, cost-effective, and easy to operate tool that can be used to non-invasively monitor plant pigment and nutrient content efficiently.
Subject(s)
Anthocyanins , Nutrients , Agriculture , Chlorophyll , CultureABSTRACT
Leaf color patterns vary depending on leaf age, pathogen infection, and environmental and nutritional stresses; thus, they are widely used to diagnose plant health statuses in agricultural fields. The visible-near infrared-shortwave infrared (VIS-NIR-SWIR) sensor measures the leaf color pattern from a wide spectral range with high spectral resolution. However, spectral information has only been employed to understand general plant health statuses (e.g., vegetation index) or phytopigment contents, rather than pinpointing defects of specific metabolic or signaling pathways in plants. Here, we report feature engineering and machine learning methods that utilize VIS-NIR-SWIR leaf reflectance for robust plant health diagnostics, pinpointing physiological alterations associated with the stress hormone, abscisic acid (ABA). Leaf reflectance spectra of wild-type, ABA2-overexpression, and deficient plants were collected under watered and drought conditions. Drought- and ABA-associated normalized reflectance indices (NRIs) were screened from all possible pairs of wavelength bands. Drought associated NRIs showed only a partial overlap with those related to ABA deficiency, but more NRIs were associated with drought due to additional spectral changes within the NIR wavelength range. Interpretable support vector machine classifiers built with 20 NRIs predicted treatment or genotype groups with an accuracy greater than those with conventional vegetation indices. Major selected NRIs were independent from leaf water content and chlorophyll content, 2 well-characterized physiological changes under drought. The screening of NRIs, streamlined with the development of simple classifiers, serves as the most efficient means of detecting reflectance bands that are highly relevant to characteristics of interest.
ABSTRACT
Gibberellins (GAs) are a class of phytohormones, important for plant growth, and very difficult to distinguish because of their similarity in chemical structures. Herein, we develop the first nanosensors for GAs by designing and engineering polymer-wrapped single-walled carbon nanotubes (SWNTs) with unique corona phases that selectively bind to bioactive GAs, GA3 and GA4, triggering near-infrared (NIR) fluorescence intensity changes. Using a new coupled Raman/NIR fluorimeter that enables self-referencing of nanosensor NIR fluorescence with its Raman G-band, we demonstrated detection of cellular GA in Arabidopsis, lettuce, and basil roots. The nanosensors reported increased endogenous GA levels in transgenic Arabidopsis mutants that overexpress GA and in emerging lateral roots. Our approach allows rapid spatiotemporal detection of GA across species. The reversible sensor captured the decreasing GA levels in salt-treated lettuce roots, which correlated remarkably with fresh weight changes. This work demonstrates the potential for nanosensors to solve longstanding problems in plant biotechnology.
Subject(s)
Arabidopsis , Nanotubes, Carbon , Gibberellins/chemistry , Gibberellins/metabolism , Plant Growth Regulators/metabolism , Arabidopsis/metabolism , Nanotubes, Carbon/chemistry , Fluorescence , Coloring AgentsABSTRACT
Calcium (Ca) deficiency causes necrotic symptoms of foliar edges known as tipburn; however, the underlying cellular mechanisms have been poorly understood due to the lack of an ideal plant model and research platform. Using a phenotyping system that quantitates growth and tipburn traits in the model bryophyte Marchantia polymorpha, we evaluated metabolic compounds and the Gß-null mutant (gpb1) that modulate the occurrence and expansion of the tipburn. Transcriptomic comparisons between wild-type and gpb1 plants revealed the phenylalanine/phenylpropanoid biosynthesis pathway and reactive oxygen species (ROS) important for Ca deficiency responses. gpb1 plants reduced ROS production possibly through transcriptomic regulations of class III peroxidases and induced the expression of phenylpropanoid pathway enzymes without changing downstream lignin contents. Supplementation of intermediate metabolites and chemical inhibitors further confirmed the cell-protective mechanisms of the phenylpropanoid and ROS pathways. Marchantia polymorpha, Arabidopsis thaliana, and Lactuca sativa showed comparable transcriptomic changes where genes related to phenylpropanoid and ROS pathways were enriched in response to Ca deficiency. In conclusion, our study demonstrated unresolved signaling and metabolic pathways of Ca deficiency response. The phenotyping platform can speed up the discovery of chemical and genetic pathways, which could be widely conserved between M. polymorpha and angiosperms.
Subject(s)
Arabidopsis , Marchantia , Calcium/metabolism , Reactive Oxygen Species/metabolism , Signal Transduction , Arabidopsis/genetics , Metabolic Networks and Pathways , GTP-Binding Proteins/metabolism , Marchantia/geneticsABSTRACT
New systems for agrochemical delivery in plants will foster precise agricultural practices and provide new tools to study plants and design crop traits, as standard spray methods suffer from elevated loss and limited access to remote plant tissues. Silk-based microneedles can circumvent these limitations by deploying a known amount of payloads directly in plants' deep tissues. However, plant response to microneedles' application and microneedles' efficacy in deploying physiologically relevant biomolecules are unknown. Here, it is shown that gene expression associated with Arabidopsis thaliana wounding response decreases within 24 h post microneedles' application. Additionally, microinjection of gibberellic acid (GA3 ) in A. thaliana mutant ft-10 provides a more effective and efficient mean than spray to activate GA3 pathways, accelerating bolting and inhibiting flower formation. Microneedle efficacy in delivering GA3 is also observed in several monocot and dicot crop species, i.e., tomato (Solanum lycopersicum), lettuce (Lactuca sativa), spinach (Spinacia oleracea), rice (Oryza Sativa), maize (Zea mays), barley (Hordeum vulgare), and soybean (Glycine max). The wide range of plants that can be successfully targeted with microinjectors opens the doors to their use in plant science and agriculture.
Subject(s)
Plants , SilkABSTRACT
The core G protein signaling module, which consists of Gα and extra-large Gα (XLG) subunits coupled with the Gßγ dimer, is a master regulator of various stress responses. In this study, we compared the basal and salt stress-induced transcriptomic, metabolomic and phenotypic profiles in Gα, Gß, and XLG-null mutants of two plant species, Arabidopsis thaliana and Marchantia polymorpha, and showed that G protein mediates the shift of transcriptional and metabolic homeostasis to stress readiness status. We demonstrated that such stress readiness serves as an intrinsic protection mechanism against further stressors through enhancing the phenylpropanoid pathway and abscisic acid responses. Furthermore, WRKY transcription factors were identified as key intermediates of G protein-mediated homeostatic shifts. Statistical and mathematical model comparisons between A. thaliana and M. polymorpha revealed evolutionary conservation of transcriptional and metabolic networks over land plant evolution, whereas divergence has occurred in the function of plant-specific atypical XLG subunit. Taken together, our results indicate that the shifts in transcriptional and metabolic homeostasis at least partially act as the mechanisms of G protein-coupled stress responses that are conserved between two distantly related plants.
Subject(s)
Arabidopsis , Marchantia , Marchantia/genetics , Arabidopsis/genetics , Metabolomics , GTP-Binding ProteinsABSTRACT
Stomata, the epidermal pores for gas exchange between plants and the atmosphere, are the major sites of water loss. During water shortage, plants limit the formation of new stoma via the phytohormone abscisic acid (ABA) to conserve water. However, how ABA suppresses stomatal production is largely unknown. Here, we demonstrate that three core SnRK2 kinases of ABA signaling inhibit the initiation and proliferation of the stomatal precursors in Arabidopsis. We show that the SnRK2s function within the precursors and directly phosphorylate SPEECHLESS (SPCH), the master transcription factor for stomatal initiation. We identify specific SPCH residues targeted by the SnRK2s, which mediate the ABA/drought-induced suppression of SPCH and stomatal production. This SnRK2-specific SPCH phosphocode connects stomatal development with ABA/drought signals and enables the independent control of this key water conservation response. Our work also highlights how distinct signaling activities can be specifically encoded on a master regulator to modulate developmental plasticity.
ABSTRACT
The copy numbers of many plant transcription factor (TF) genes substantially increased during terrestrialization. This allowed TFs to acquire new specificities and thus create gene regulatory networks (GRNs) with new biological functions to help plants adapt to terrestrial environments. Through characterizing heat shock factor (HSF) genes MpHSFA1 and MpHSFB1 in the liverwort Marchantia polymorpha, we explored how heat-responsive GRNs widened their functions in M. polymorpha and Arabidopsis thaliana. An interspecies comparison of heat-induced transcriptomes and the evolutionary rates of HSFs demonstrated the emergence and subsequent rapid evolution of HSFB prior to terrestrialization. Transcriptome and metabolome analyses of M. polymorpha HSF-null mutants revealed that MpHSFA1 controls canonical heat responses such as thermotolerance and metabolic changes. MpHSFB1 also plays essential roles in heat responses, as well as regulating developmental processes including meristem branching and antheridiophore formation. Analysis of cis-regulatory elements revealed development- and stress-related TFs that function directly or indirectly downstream of HSFB. Male gametophytes of M. polymorpha showed higher levels of thermotolerance than female gametophytes, which could be explained by different expression levels of MpHSFA1U and MpHSFA1V on sex chromosome. We propose that the diversification of HSFs is linked to the expansion of HS responses, which enabled coordinated multicellular reactions in land plants.
Subject(s)
Arabidopsis , Marchantia , Arabidopsis/metabolism , Gene Expression Regulation, Plant/genetics , Gene Regulatory Networks , Heat Shock Transcription Factors/metabolism , Heat-Shock Response/genetics , Marchantia/genetics , Marchantia/metabolism , Plant Proteins/genetics , Plant Proteins/metabolismABSTRACT
Different environmental stresses often evoke similar physiological disorders such as growth retardation; however, specific consequences reported among individual stresses indicate potential mechanisms to distinguish different stress types in plants. Here, we examined mechanisms to differentiate between stress types in Arabidopsis (Arabidopsis thaliana). Gene expression patterns recapitulating several abiotic stress responses suggested abscisic acid (ABA) as a mediator of the common stress response, while stress type-specific responses were related to metabolic adaptations. Transcriptome and metabolome analyses identified Arabidopsis Gß (AGB1) mediating the common stress-responsive genes and primary metabolisms under nitrogen excess. AGB1 regulated the expressions of multiple WRKY transcription factors. Gene Ontology and mutant analyses revealed different roles among WRKYs: WRKY40 is involved in ABA and common stress responses, while WRKY75 regulates metabolic processes. The AGB1-WRKY signaling module controlled developmental plasticity in roots under nitrogen excess. Signal transmission from AGB1 to a selective set of WRKYs would be essential to evoke unique responses to different types of stresses.
Subject(s)
Arabidopsis Proteins , Arabidopsis , GTP-Binding Protein beta Subunits , Abscisic Acid/metabolism , Abscisic Acid/pharmacology , Arabidopsis/metabolism , Arabidopsis Proteins/metabolism , GTP-Binding Protein beta Subunits/genetics , Gene Expression Regulation, Plant , Nitrogen/metabolism , Stress, Physiological/genetics , Transcription Factors/metabolismABSTRACT
Fluorescence imaging has shown great potential in non-invasive plant monitoring and analysis. However, current systems have several limitations, such as bulky size, high cost, contact measurement, and lack of multifunctionality, which may hinder its applications in a wide range of settings including indoor vertical farming. Herein, we developed a compact handheld fluorescence imager enabling multipurpose plant phenotyping, such as continuous photosynthetic activity monitoring and non-destructive anthocyanin quantification. The compact imager comprises of pulse-amplitude-modulated multi-color light emitting diodes (LEDs), optimized light illumination and collection, dedicated driver circuit board, miniaturized charge-coupled device camera, and associated image analytics. Experiments conducted in drought stressed lettuce proved that the novel imager could quantitatively evaluate the plant stress by the non-invasive measurement of photosynthetic activity efficiency. Moreover, a non-invasive and fast quantification of anthocyanins in green and red Batavia lettuce leaves had excellent correlation (>84%) with conventional destructive biochemical analysis. Preliminary experimental results emphasize the high throughput monitoring capability and multifunctionality of our novel handheld fluorescence imager, indicating its tremendous potential in modern agriculture.
ABSTRACT
Plant cells constantly alter their gene expression profiles to respond to environmental fluctuations. These continuous adjustments are regulated by multi-hierarchical networks of transcription factors. To understand how such gene regulatory networks (GRNs) have stabilized evolutionarily while allowing for species-specific responses, we compare the GRNs underlying salt response in the early-diverging and late-diverging plants Marchantia polymorpha and Arabidopsis thaliana. Salt-responsive GRNs, constructed on the basis of the temporal transcriptional patterns in the two species, share common trans-regulators but exhibit an evolutionary divergence in cis-regulatory sequences and in the overall network sizes. In both species, WRKY-family transcription factors and their feedback loops serve as central nodes in salt-responsive GRNs. The divergent cis-regulatory sequences of WRKY-target genes are probably associated with the expansion in network size, linking salt stress to tissue-specific developmental and physiological responses. The WRKY modules and highly linked WRKY feedback loops have been preserved widely in other plants, including rice, while keeping their binding-motif sequences mutable. Together, the conserved trans-regulators and the quickly evolving cis-regulatory sequences allow salt-responsive GRNs to adapt over a long evolutionary timescale while maintaining some consistent regulatory structure. This strategy may benefit plants as they adapt to changing environments.
Subject(s)
Arabidopsis/genetics , Gene Regulatory Networks , Marchantia/genetics , Plant Proteins/genetics , Salt Stress/genetics , Adaptation, Physiological , Arabidopsis Proteins/genetics , Biological Evolution , Gene Expression Regulation, Plant , Mutation , Oryza/genetics , Phylogeny , Transcription Factors/geneticsABSTRACT
The extra-large guanosine-5'-triphosphate (GTP)-binding protein 2, XLG2, is an unconventional Gα subunit of the Arabidopsis (Arabidopsis thaliana) heterotrimeric GTP-binding protein complex with a major role in plant defense. In vitro biochemical analyses and molecular dynamic simulations show that affinity of XLG2 for GTP is two orders of magnitude lower than that of the conventional Gα, AtGPA1. Here we tested the physiological relevance of GTP binding by XLG2. We generated an XLG2(T476N) variant with abolished GTP binding, as confirmed by in vitro GTPγS binding assay. Yeast three-hybrid, bimolecular fluorescence complementation, and split firefly-luciferase complementation assays revealed that the nucleotide-depleted XLG2(T476N) retained wild-type XLG2-like interactions with the Gßγ dimer and defense-related receptor-like kinases. Both wild-type and nucleotide-depleted XLG2(T476N) restored the defense responses against Fusarium oxysporum and Pseudomonas syringae compromised in the xlg2 xlg3 double mutant. Additionally, XLG2(T476N) was fully functional restoring stomatal density, root growth, and sensitivity to NaCl, but failed to complement impaired germination and vernalization-induced flowering. We conclude that XLG2 is able to function in a GTP-independent manner and discuss its possible mechanisms of action.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/genetics , Fusarium/physiology , Heterotrimeric GTP-Binding Proteins/metabolism , Plant Diseases/immunology , Pseudomonas syringae/physiology , Arabidopsis/enzymology , Arabidopsis/immunology , Arabidopsis/microbiology , Arabidopsis Proteins/genetics , Guanosine Triphosphate/metabolism , Heterotrimeric GTP-Binding Proteins/genetics , Plant Diseases/microbiologyABSTRACT
Plant growth and grain filling are the key agronomical traits for grain weight and yield of rice. The continuous improvement in rice yield is required for a future sustainable global economy and food security. The heterotrimeric G protein complex containing a canonical α subunit (RGA1) couples extracellular signals perceived by receptors to modulate cell function including plant development and grain weight. We hypothesized that, besides RGA1, three atypical, extra-large GTP-binding protein (XLG) subunits also regulate panicle architecture, plant growth, development, grain weight, and disease resistance. Here, we identified a role of XLGs in agronomic traits and stress tolerance by genetically ablating all three rice XLGs individually and in combination using the CRISPR/Cas9 genome editing in rice. For this study, eight (three single, two double, and three triple) null mutants were selected. Three XLG proteins combinatorically regulate seed filling, because loss confers a decrease in grain weight from 14% with loss of one XLG and loss of three to 32% decrease in grain weight. Null mutations in XLG2 and XLG4 increase grain size. The mutants showed significantly reduced panicle length and number per plant including lesser number of grains per panicle compared to the controls. Loss-of-function of all individual XLGs contributed to 9% more aerial biomass compared to wild type (WT). The double mutant showed improved salinity tolerance. Moreover, loss of the XLG gene family confers hypersensitivity to pathogens. Our findings suggest that the non-canonical XLGs play important roles in regulating rice plant growth, grain filling, panicle phenotype, stress tolerance, and disease resistance. Genetic manipulation of XLGs has the potential to improve agronomic properties in rice.
ABSTRACT
Nutrient stresses induce foliar chlorosis and growth defects. Here we propose heterotrimeric G proteins as signaling mediators of various nutrient stresses, through meta-analyses of >20 transcriptomic data sets associated with nutrient stresses or G protein mutations. Systematic comparison of transcriptomic data yielded 104 genes regulated by G protein subunits under common nutrient stresses: 69 genes under Gß subunit (AGB1) control and 35 genes under Gα subunit (GPA1) control. Quantitative real-time PCR experiments validate that several transcription factors and metal transporters changed in expression level under suboptimal iron, zinc, and/or copper concentrations, while being misregulated in the Arabidopsis Gß-null (agb1) mutant. The agb1 mutant had altered metal ion profiles and exhibited severe growth arrest under zinc stress, and aberrant root waving under iron and zinc stresses, while the Gα-null mutation attenuated leaf chlorosis under iron deficiency in both Arabidopsis and rice. Our transcriptional network analysis inferred computationally that WRKY-family transcription factors mediate the AGB1-dependent nutrient responses. As corroborating evidence of our inference, ectopic expression of WRKY25 or WRKY33 rescued the zinc stress phenotypes and the expression of zinc transporters in the agb1-2 background. These results, together with Gene Ontology analyses, suggest two contrasting roles for G protein-coupled signaling pathways in micronutrient stress responses: one enhancing general stress tolerance and the other modulating ion homeostasis through WRKY transcriptional regulatory networks. In addition, tolerance to iron stress in the rice Gα mutant provides an inroad to improve nutrient stress tolerance of agricultural crops by manipulating G protein signaling.
Subject(s)
Arabidopsis Proteins , GTP-Binding Protein beta Subunits , Heterotrimeric GTP-Binding Proteins , Arabidopsis Proteins/metabolism , GTP-Binding Protein beta Subunits/genetics , GTP-Binding Protein beta Subunits/metabolism , Gene Expression Regulation, Plant , Heterotrimeric GTP-Binding Proteins/genetics , Micronutrients , Transcription Factors/geneticsABSTRACT
The heterotrimeric G protein complex, composed of Gα, Gß, and Gγ subunits, plays some role in structural development in plants but this role could be indirect because loss-of-function mutations do not alter the body plan and post-embryonic organs differ only morphologically and not in their identity. This uncertainty has been compounded by the fact that loss of the Gß subunit in cereals, but not Arabidopsis, is seedling lethal and that loss of maize Gα subunit confers prolificacy of a reproductive organ. In this study, we comprehensively profiled the root and shoot structural traits of rice Gα-null and viable Gß-RNAi "knockdown" mutants, and found anomalous morphologies caused by Gß-RNAi that are distinct from the Arabidopsis orthologue. The rice Gß-RNAi mutant exhibited reduced radial growth of aerial parts as well as a more compact root architecture, among which smaller root mass seems mainly due to increased necrosis when grown on soil. In addition, three dimensional analyses of rice root system architecture revealed that the smaller root architecture of Gß-RNAi plant is also due to both reduced root elongation and adventitious root formation. This contrasts to the Arabidopsis Gß-null mutation that promotes cell proliferation. There is elevated cell senescence activity both visualized by Evans Blue staining and inferred from an expression analysis of cell-death marker genes. We propose that the morphological phenotypes of rice Gß-RNAi plants are predominantly associated with the mediation of various stresses and cell senescence, consistent with an indirect role for Arabidopsis Gß in development where the orthologous gene ablation mainly confers altered cell proliferation. We also elaborate our speculative working hypothesis that cell division is a type of stress and as such due to impairment in responding to stress in the G protein mutants, manifests as altered morphology and architecture but not an altered body plan or organ identities.