RESUMO
In plants so-called plasma membrane intrinsic proteins (PIPs) are major water channels governing plant water status. Membrane trafficking contributes to functional regulation of major PIPs and is crucial for abiotic stress resilience. Arabidopsis PIP2;1 is rapidly internalised from the plasma membrane in response to high salinity to regulate osmotic water transport, but knowledge of the underlying mechanisms is fragmentary. Here we show that PIP2;1 occurs in complex with SYNTAXIN OF PLANTS 132 (SYP132) together with the plasma membrane H+-ATPase AHA1 as evidenced through in vivo and in vitro analysis. SYP132 is a multifaceted vesicle trafficking protein, known to interact with AHA1 and promote endocytosis to impact growth and pathogen defence. Tracking native proteins in immunoblot analysis, we found that salinity stress enhances SYP132 interactions with PIP2;1 and PIP2;2 isoforms to promote redistribution of the water channels away from the plasma membrane. Concurrently, AHA1 binding within the SYP132-complex was significantly reduced under salinity stress and increased the density of AHA1 proteins at the plasma membrane in leaf tissue. Manipulating SYP132 function in Arabidopsis thaliana enhanced resilience to salinity stress and analysis in heterologous systems suggested that the SNARE influences PIP2;1 osmotic water permeability. We propose therefore that SYP132 coordinates AHA1 and PIP2;1 abundance at the plasma membrane and influences leaf hydraulics to regulate plant responses to abiotic stress signals.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Proteínas Qa-SNARE , Estresse Salino , Aquaporinas/metabolismo , Aquaporinas/genética , Arabidopsis/fisiologia , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Membrana Celular/metabolismo , Transporte Proteico , ATPases Translocadoras de Prótons/metabolismo , ATPases Translocadoras de Prótons/genética , Proteínas Qa-SNARE/metabolismo , Proteínas Qa-SNARE/genética , Proteínas SNARE/metabolismo , Proteínas SNARE/genéticaRESUMO
If the past century marked the birth of membrane transport as a focus for research in plants, the past 50 years has seen the field mature from arcane interest to a central pillar of plant physiology. Ion transport across plant membranes accounts for roughly 30% of the metabolic energy consumed by a plant cell, and it underpins virtually every aspect of plant biology, from mineral nutrition, cell expansion, and development to auxin polarity, fertilization, plant pathogen defense, and senescence. The means to quantify ion flux through individual transporters, even single channel proteins, became widely available as voltage clamp methods expanded from giant algal cells to the fungus Neurospora crassa in the 1970s and the cells of angiosperms in the 1980s. Here, I touch briefly on some key aspects of the development of modern electrophysiology with a focus on the guard cells of stomata, now without dispute the premier plant cell model for ion transport and its regulation. Guard cells have proven to be a crucible for many technical and conceptual developments that have since emerged into the mainstream of plant science. Their study continues to provide fundamental insights and carries much importance for the global challenges that face us today.
Assuntos
Transporte de Íons , Estômatos de Plantas , Plantas , Plantas/metabolismo , Estômatos de Plantas/metabolismo , Estômatos de Plantas/fisiologia , Membrana Celular/metabolismoRESUMO
The evolution of stomata marks one of the key advances that enabled plants to colonise dry land while allowing gas exchange for photosynthesis. In large measure, stomata retain a common design across species that incorporates paired guard cells with little variation in structure. By contrast, the cells of the stomatal complex immediately surrounding the guard cells vary widely in shape, size and count. Their origins in development are similarly diverse. Thus, the surrounding cells are likely a luxury that the necessity of stomatal control cannot do without (with apologies to Oscar Wilde). Surrounding cells are thought to support stomatal movements as solute reservoirs and to shape stomatal kinetics through backpressure on the guard cells. Their variety may also reflect a substantial diversity in function. Certainly modelling, kinetic analysis and the few electrophysiological studies to date give hints of much more complex contributions in stomatal physiology. Even so, our knowledge of the cells surrounding the guard cells in the stomatal complex is far from complete.
Assuntos
Estômatos de Plantas , Estômatos de Plantas/fisiologia , Evolução Biológica , Modelos Biológicos , Fotossíntese/fisiologiaRESUMO
Stomata are microscopic pores at the surface of plant leaves that facilitate gaseous diffusion to support photosynthesis. The guard cells around each stoma regulate the pore aperture. Plants that carry out C4 photosynthesis are usually more resilient than C3 plants to stress, and their stomata operate over a lower dynamic range of CO2 within the leaf. What makes guard cells of C4 plants more responsive than those of C3 plants? We used gas exchange and electrophysiology, comparing stomatal kinetics of the C4 plant Gynandropsis gynandra and the phylogenetically related C3 plant Arabidopsis thaliana. We found, with varying CO2 and light, that Gynandropsis showed faster changes in stomata conductance and greater water use efficiency when compared with Arabidopsis. Electrophysiological analysis of the dominant K+ channels showed that the outward-rectifying channels, responsible for K+ loss during stomatal closing, were characterised by a greater maximum conductance and substantial negative shift in the voltage dependence of gating, indicating a reduced inhibition by extracellular K+ and enhanced capacity for K+ flux. These differences correlated with the accelerated stomata kinetics of Gynandropsis, suggesting that subtle changes in the biophysical properties of a key transporter may prove a target for future efforts to engineer C4 stomatal kinetics.
Assuntos
Arabidopsis , Magnoliopsida , Estômatos de Plantas/fisiologia , Dióxido de Carbono , Folhas de Planta/fisiologia , Fotossíntese/fisiologia , Arabidopsis/fisiologia , GasesRESUMO
Stomata play a crucial role in plants by controlling water status and responding to drought stress. However, simultaneously improving stomatal opening and drought tolerance has proven to be a significant challenge. To address this issue, we employed the OnGuard quantitative model, which accurately represents the mechanics and coordination of ion transporters in guard cells. With the guidance of OnGuard, we successfully engineered plants that overexpressed the main tonoplast Ca2+-ATPase gene, ACA11, which promotes stomatal opening and enhances plant growth. Surprisingly, these transgenic plants also exhibited improved drought tolerance due to reduced water loss through their stomata. Again, OnGuard assisted us in understanding the mechanism behind the unexpected stomatal behaviors observed in the ACA11 overexpressing plants. Our study revealed that the overexpression of ACA11 facilitated the accumulation of Ca2+ in the vacuole, thereby influencing Ca2+ storage and leading to an enhanced Ca2+ elevation in response to abscisic acid. This regulatory cascade finely tunes stomatal responses, ultimately leading to enhanced drought tolerance. Our findings underscore the importance of tonoplast Ca2+-ATPase in manipulating stomatal behavior and improving drought tolerance. Furthermore, these results highlight the diverse functions of tonoplast-localized ACA11 in response to different conditions, emphasizing its potential for future applications in plant enhancement.
Assuntos
ATPases Transportadoras de Cálcio , Resistência à Seca , Estômatos de Plantas , Plantas Geneticamente Modificadas , Ácido Abscísico/farmacologia , Ácido Abscísico/metabolismo , Arabidopsis/genética , Arabidopsis/fisiologia , Cálcio/metabolismo , ATPases Transportadoras de Cálcio/metabolismo , ATPases Transportadoras de Cálcio/genética , Resistência à Seca/genética , Regulação da Expressão Gênica de Plantas , Estômatos de Plantas/fisiologia , Estômatos de Plantas/genética , Vacúolos/metabolismoRESUMO
Gas exchange across the stomatal pores of leaves is a focal point in studies of plant-environmental relations. Stomata regulate atmospheric exchange with the inner air spaces of the leaf. They open to allow CO2 entry for photosynthesis and close to minimize water loss. Models that focus on the phenomenology of stomatal conductance generally omit the mechanics of the guard cells that regulate the pore aperture. The OnGuard platform fills this gap and offers a truly mechanistic approach with which to analyse stomatal gas exchange, whole-plant carbon assimilation and water-use efficiency. Previously, OnGuard required specialist knowledge of membrane transport, signalling and metabolism. Here we introduce OnGuard3e, a software package accessible to ecophysiologists and membrane biologists alike. We provide a brief guide to its use and illustrate how the package can be applied to explore and analyse stomatal conductance, assimilation and water use efficiencies, addressing a range of experimental questions with truly predictive outputs.
Assuntos
Folhas de Planta , Estômatos de Plantas , Estômatos de Plantas/fisiologia , Folhas de Planta/metabolismo , Fotossíntese/fisiologia , Plantas/metabolismo , Água/metabolismo , Dióxido de Carbono/metabolismoRESUMO
Vesicle exocytosis underpins signaling and development in plants and is vital for cell expansion. Vesicle tethering and fusion are thought to occur sequentially, with tethering mediated by the exocyst and fusion driven by assembly of soluble NSF attachment protein receptor (SNARE) proteins from the vesicle membrane (R-SNAREs or vesicle-associated membrane proteins [VAMPs]) and the target membrane (Q-SNAREs). Interactions between exocyst and SNARE protein complexes are known, but their functional consequences remain largely unexplored. We now identify a hierarchy of interactions leading to secretion in Arabidopsis (Arabidopsis thaliana). Mating-based split-ubiquitin screens and in vivo Förster resonance energy transfer analyses showed that exocyst EXO70 subunits bind preferentially to cognate plasma membrane SNAREs, notably SYP121 and VAMP721. The exo70A1 mutant affected SNARE distribution and suppressed vesicle traffic similarly to the dominant-negative truncated protein SYP121ΔC, which blocks secretion at the plasma membrane. These phenotypes are consistent with the epistasis of exo70A1 in the exo70A1 syp121 double mutant, which shows decreased growth similar to exo70A1 single mutants. However, the exo70A1 vamp721 mutant showed a strong, synergy, suppressing growth and cell expansion beyond the phenotypic sum of the two single mutants. These data are best explained by a hierarchy of SNARE recruitment to the exocyst at the plasma membrane, dominated by the R-SNARE and plausibly with the VAMP721 longin domain as a nexus for binding.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Proteínas SNARE/metabolismo , Arabidopsis/citologia , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Membrana Celular/metabolismo , Exocitose/fisiologia , Transferência Ressonante de Energia de Fluorescência , Mutação , Plantas Geneticamente Modificadas , Proteínas Qa-SNARE/genética , Proteínas Qa-SNARE/metabolismo , Proteínas R-SNARE/genética , Proteínas R-SNARE/metabolismo , Proteínas SNARE/genéticaRESUMO
Starch in Arabidopsis (Arabidopsis thaliana) guard cells is rapidly degraded at the start of the day by the glucan hydrolases α-AMYLASE3 (AMY3) and ß-AMYLASE1 (BAM1) to promote stomatal opening. This process is activated via phototropin-mediated blue light signaling downstream of the plasma membrane H+-ATPase. It remains unknown how guard cell starch degradation integrates with light-regulated membrane transport processes in the fine control of stomatal opening kinetics. We report that H+, K+, and Cl- transport across the guard cell plasma membrane is unaltered in the amy3 bam1 mutant, suggesting that starch degradation products do not directly affect the capacity to transport ions. Enzymatic quantification revealed that after 30 min of blue light illumination, amy3 bam1 guard cells had similar malate levels as the wild type, but had dramatically altered sugar homeostasis, with almost undetectable amounts of Glc. Thus, Glc, not malate, is the major starch-derived metabolite in Arabidopsis guard cells. We further show that impaired starch degradation in the amy3 bam1 mutant resulted in an increase in the time constant for opening of 40 min. We conclude that rapid starch degradation at dawn is required to maintain the cytoplasmic sugar pool, clearly needed for fast stomatal opening. The conversion and exchange of metabolites between subcellular compartments therefore coordinates the energetic and metabolic status of the cell with membrane ion transport.
Assuntos
Arabidopsis/citologia , Arabidopsis/fisiologia , Glucose/metabolismo , Estômatos de Plantas/fisiologia , Amido/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Transporte Biológico , Cloretos/metabolismo , Escuridão , Luz , Malatos/metabolismo , Mutação , Fotossíntese , Células Vegetais/metabolismo , Potássio/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , ATPases Translocadoras de Prótons/genética , ATPases Translocadoras de Prótons/metabolismo , PrótonsRESUMO
Membrane voltage arises from the transport of ions through ion-translocating ATPases, ion-coupled transport of solutes, and ion channels, and is an integral part of the bioenergetic "currency" of the membrane. The dynamics of membrane voltage-so-called action, systemic, and variation potentials-have also led to a recognition of their contributions to signal transduction, both within cells and across tissues. Here, we review the origins of our understanding of membrane voltage and its place as a central element in regulating transport and signal transmission. We stress the importance of understanding voltage as a common intermediate that acts both as a driving force for transport-an electrical "substrate"-and as a product of charge flux across the membrane, thereby interconnecting all charge-carrying transport across the membrane. The voltage interconnection is vital to signaling via second messengers that rely on ion flux, including cytosolic free Ca2+, H+, and the synthesis of reactive oxygen species generated by integral membrane, respiratory burst oxidases. These characteristics inform on the ways in which long-distance voltage signals and voltage oscillations give rise to unique gene expression patterns and influence physiological, developmental, and adaptive responses such as systemic acquired resistance to pathogens and to insect herbivory.
Assuntos
Transporte Biológico/fisiologia , Membrana Celular/fisiologia , Transporte de Íons/fisiologia , Desenvolvimento Vegetal , Transdução de Sinais/fisiologia , Canais de Ânion Dependentes de Voltagem/fisiologiaRESUMO
Activation of plasma membrane (PM) H+-ATPase activity is crucial in guard cells to promote light-stimulated stomatal opening, and in growing organs to promote cell expansion. In growing organs, SMALL AUXIN UP RNA (SAUR) proteins inhibit the PP2C.D2, PP2C.D5, and PP2C.D6 (PP2C.D2/5/6) phosphatases, thereby preventing dephosphorylation of the penultimate phosphothreonine of PM H+-ATPases and trapping them in the activated state to promote cell expansion. To elucidate whether SAUR-PP2C.D regulatory modules also affect reversible cell expansion, we examined stomatal apertures and conductances of Arabidopsis thaliana plants with altered SAUR or PP2C.D activity. Here, we report that the pp2c.d2/5/6 triple knockout mutant plants and plant lines overexpressing SAUR fusion proteins exhibit enhanced stomatal apertures and conductances. Reciprocally, saur56 saur60 double mutants, lacking two SAUR genes normally expressed in guard cells, displayed reduced apertures and conductances, as did plants overexpressing PP2C.D5. Although altered PM H+-ATPase activity contributes to these stomatal phenotypes, voltage clamp analysis showed significant changes also in K+ channel gating in lines with altered SAUR and PP2C.D function. Together, our findings demonstrate that SAUR and PP2C.D proteins act antagonistically to facilitate stomatal movements through a concerted targeting of both ATP-dependent H+ pumping and channel-mediated K+ transport.
Assuntos
Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Membrana Celular/metabolismo , Ácidos Indolacéticos/metabolismo , Monoéster Fosfórico Hidrolases/metabolismo , Estômatos de Plantas/metabolismo , ATPases Translocadoras de Prótons/metabolismo , Ecótipo , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Variação Genética , Reguladores de Crescimento de Plantas/metabolismoRESUMO
Chloroplast retrograde signaling networks are vital for chloroplast biogenesis, operation, and signaling, including excess light and drought stress signaling. To date, retrograde signaling has been considered in the context of land plant adaptation, but not regarding the origin and evolution of signaling cascades linking chloroplast function to stomatal regulation. We show that key elements of the chloroplast retrograde signaling process, the nucleotide phosphatase (SAL1) and 3'-phosphoadenosine-5'-phosphate (PAP) metabolism, evolved in streptophyte algae-the algal ancestors of land plants. We discover an early evolution of SAL1-PAP chloroplast retrograde signaling in stomatal regulation based on conserved gene and protein structure, function, and enzyme activity and transit peptides of SAL1s in species including flowering plants, the fern Ceratopteris richardii, and the moss Physcomitrella patens Moreover, we demonstrate that PAP regulates stomatal closure via secondary messengers and ion transport in guard cells of these diverse lineages. The origin of stomata facilitated gas exchange in the earliest land plants. Our findings suggest that the conquest of land by plants was enabled by rapid response to drought stress through the deployment of an ancestral SAL1-PAP signaling pathway, intersecting with the core abscisic acid signaling in stomatal guard cells.
Assuntos
Adaptação Fisiológica , Evolução Biológica , Cloroplastos/metabolismo , Transdução de Sinais , Viridiplantae/fisiologia , Difosfato de Adenosina , Embriófitas/fisiologia , Peróxido de Hidrogênio/metabolismo , Transporte de Íons , Movimento , Óxido Nítrico/metabolismo , Filogenia , Estômatos de Plantas/fisiologiaRESUMO
It has been proposed by some plant scientists that plants are cognitive and conscious organisms, although this is a minority view. Here we present a brief summary of some of the arguments against this view, followed by a critique of an article in this same issue of Biochemical and Biophysical Research Communications by Calvo, Baluska, and Trewavas (2020) that cites Integrated Information Theory (IIT) as providing additional support for plant consciousness. The authors base their argument on the assumptions that all cells are conscious and that consciousness is confined to life. However, IIT allows for consciousness in various nonliving systems, and thus does not restrict consciousness to living organisms. Therefore, IIT cannot be used to prove plant consciousness, for which there is neither empirical evidence nor support from other, neuron-based, theories of consciousness.
Assuntos
Estado de Consciência/fisiologia , Teoria da Informação , Plantas/metabolismo , HumanosRESUMO
Ferns appear in the fossil record some 200 Myr before angiosperms. However, as angiosperm-dominated forest canopies emerged in the Cretaceous period there was an explosive diversification of modern (leptosporangiate) ferns, which thrived in low, blue-enhanced light beneath angiosperm canopies. A mechanistic explanation for this transformative event in the diversification of ferns has remained elusive. We used physiological assays, transcriptome analysis and evolutionary bioinformatics to investigate a potential connection between the evolution of enhanced stomatal sensitivity to blue light in modern ferns and the rise of angiosperm-dominated forests in the geological record. We demonstrate that members of the largest subclade of leptosporangiate ferns, Polypodiales, have significantly faster stomatal response to blue light than more ancient fern lineages and a representative angiosperm. We link this higher sensitivity to levels of differentially expressed genes in blue-light signaling, particularly in the cryptochrome (CRY) signaling pathway. Moreover, CRYs of the Polypodiales examined show gene duplication events between 212.9-196.9 and 164.4-151.8 Ma, when angiosperms were emerging, which are lacking in other major clades of extant land plants. These findings suggest that evolution of stomatal blue-light sensitivity helped modern ferns exploit the shady habitat beneath angiosperm forest canopies, fueling their Cretaceous hyperdiversification.
Assuntos
Substâncias Explosivas , Gleiquênias , Magnoliopsida , Evolução Biológica , Gleiquênias/genética , Florestas , Fósseis , Magnoliopsida/genética , FilogeniaRESUMO
Plants are known to exhibit a thigmomorphogenetic response to mechanical stimuli by altering their morphology and mechanical properties. Wind is widely perceived as mechanical stress and in many experiments its influence is simulated by applying mechanical perturbations. However, it is known that wind-induced effects on plants can differ and at times occur even in the opposite direction compared with those induced by mechanical perturbations. In the present study, the long-term response of Arabidopsis thaliana to a constant unidirectional wind was investigated. We found that exposure to wind resulted in a positive anemotropic response and in significant alterations to Arabidopsis morphology, mechanical properties, and anatomical tissue organization that were associated with the plant's strategy of acclimation to a windy environment. Overall, the observed response of Arabidopsis to wind differs significantly from previously reported responses of Arabidopsis to mechanical perturbations. The presented results suggest that the response of Arabidopsis is sensitive to the type of mechanical stimulus applied, and that it is not always straightforward to simulate one type of perturbation by another.
Assuntos
Arabidopsis , Estresse Mecânico , VentoRESUMO
Plant membrane transport, like transport across all eukaryotic membranes, is highly non-linear and leads to interactions with characteristics so complex that they defy intuitive understanding. The physiological behaviour of stomatal guard cells is a case in point in which, for example, mutations expected to influence stomatal closing have profound effects on stomatal opening and manipulating transport across the vacuolar membrane affects the plasma membrane. Quantitative mathematical modelling is an essential tool in these circumstances, both to integrate the knowledge of each transport process and to understand the consequences of their manipulation in vivo. Here, we outline the OnGuard modelling environment and its use as a guide to predicting the emergent properties arising from the interactions between non-linear transport processes. We summarise some of the recent insights arising from OnGuard, demonstrate its utility in interpreting stomatal behaviour, and suggest ways in which the OnGuard environment may facilitate 'reverse-engineering' of stomata to improve water use efficiency and carbon assimilation.
Assuntos
Arabidopsis/fisiologia , Membrana Celular/fisiologia , Estômatos de Plantas/fisiologia , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/fisiologia , Transporte Biológico , Carbono/metabolismo , Engenharia Genética , Cinética , Modelos Teóricos , Mutação , Osmose , Folhas de Planta/fisiologia , ATPases Translocadoras de Prótons/genética , ATPases Translocadoras de Prótons/fisiologia , Vacúolos/fisiologia , Água/fisiologiaRESUMO
Plants utilising crassulacean acid metabolism (CAM) concentrate CO2 around RuBisCO while reducing transpirational water loss associated with photosynthesis. Unlike stomata of C3 and C4 species, CAM stomata open at night for the mesophyll to fix CO2 into malate (Mal) and store it in the vacuole. CAM plants decarboxylate Mal in the light, generating high CO2 concentrations within the leaf behind closed stomata for refixation by RuBisCO. CO2 may contribute to stomatal closure but additional mechanisms, plausibly including Mal activation of anion channels, ensure closure in the light. In the CAM species Kalanchoë fedtschenkoi, we found that guard cell anion channel activity, recorded under voltage clamp, follows KfSLAC1 and KfALMT12 transcript abundance, declining to near zero by the end of the light period. Unexpectedly, however, we found that extracellular Mal inhibited the anion current of Kalanchoë guard cells, both in wild-type and RNAi mutants with impaired Mal metabolism. We conclude that the diurnal cycle of anion channel gene transcription, rather than the physiological signal of Mal release, is a key factor in the inverted CAM stomatal cycle.
Assuntos
Kalanchoe , Malatos , Ânions , Metabolismo Ácido das Crassuláceas , FotossínteseRESUMO
SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) proteins facilitate vesicle traffic through their assembly in a heteromeric complex that drives membrane fusion. Much of vesicle traffic at the Arabidopsis (Arabidopsis thaliana) plasma membrane is subject to the Sec1/Munc18 protein SEC11, which, along with plasma membrane K+ channels, selectively binds with the SNARE SYP121 to regulate its assembly in complex. How SEC11 binding is coordinated with the K+ channels is poorly understood, as both SEC11 and the channels are thought to compete for the same SNARE binding site. Here, we identify a second binding motif within the N terminus of SYP121 and demonstrate that this motif affects SEC11 binding independently of the F9xRF motif that is shared with the K+ channels. This second, previously unrecognized motif is centered on residues R20R21 of SYP121 and is essential for SEC11 interaction with SYP121. Mutation of the R20R21 motif blocked vesicle traffic without uncoupling the effects of SYP121 on solute and K+ uptake associated with the F9xRF motif; the mutation also mimicked the effects on traffic block observed on coexpression of the dominant-negative SEC11Δ149 fragment. We conclude that the R20R21 motif represents a secondary site of interaction for the Sec1/Munc18 protein during the transition of SYP121 from the occluded to the open conformation that leads to SNARE complex assembly.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas Qa-SNARE/metabolismo , Motivos de Aminoácidos , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/genética , Sítios de Ligação , Proteínas de Ciclo Celular/genética , Mutação , Plantas Geneticamente Modificadas , Canais de Potássio/metabolismo , Domínios e Motivos de Interação entre Proteínas , Proteínas SNARE/genética , Proteínas SNARE/metabolismoRESUMO
Cell expansion requires that ion transport and secretory membrane traffic operate in concert. Evidence from Arabidopsis (Arabidopsis thaliana) indicates that such coordination is mediated by physical interactions between subsets of so-called SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) proteins, which drive the final stages of vesicle fusion, and K+ channels, which facilitate uptake of the cation to maintain cell turgor pressure as the cell expands. However, the sequence of SNARE binding with the K+ channels and its interweaving within the events of SNARE complex assembly for exocytosis remains unclear. We have combined protein-protein interaction and electrophysiological analyses to resolve the binding interactions of the hetero-oligomeric associations. We find that the RYxxWE motif, located within the voltage sensor of the K+ channels, is a nexus for multiple SNARE interactions. Of these, K+ channel binding and its displacement of the regulatory protein SEC11 is critical to prime the Qa-SNARE SYP121. Our results indicate a stabilizing role for the Qbc-SNARE SNAP33 in the Qa-SNARE transition to SNARE complex assembly with the R-SNARE VAMP721. They also suggest that, on its own, the R-SNARE enters an anomalous binding mode with the channels, possibly as a fail-safe measure to ensure a correct binding sequence. Thus, we suggest that SYP121 binding to the K+ channels serves the role of a primary trigger to initiate assembly of the secretory machinery for exocytosis.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Proteínas de Ciclo Celular/metabolismo , Canais de Potássio/metabolismo , Proteínas SNARE/metabolismo , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Cátions/metabolismo , Proteínas de Ciclo Celular/genética , Membrana Celular/metabolismo , Exocitose , Canais de Potássio/genética , Transporte Proteico , Proteínas Qa-SNARE/genética , Proteínas Qa-SNARE/metabolismo , Proteínas SNARE/genéticaRESUMO
Stomatal movements depend on the transport and metabolism of osmotic solutes that drive reversible changes in guard cell volume and turgor. These processes are defined by a deep knowledge of the identities of the key transporters and of their biophysical and regulatory properties, and have been modeled successfully with quantitative kinetic detail at the cellular level. Transpiration of the leaf and canopy, by contrast, is described by quasilinear, empirical relations for the inputs of atmospheric humidity, CO2, and light, but without connection to guard cell mechanics. Until now, no framework has been available to bridge this gap and provide an understanding of their connections. Here, we introduce OnGuard2, a quantitative systems platform that utilizes the molecular mechanics of ion transport, metabolism, and signaling of the guard cell to define the water relations and transpiration of the leaf. We show that OnGuard2 faithfully reproduces the kinetics of stomatal conductance in Arabidopsis thaliana and its dependence on vapor pressure difference (VPD) and on water feed to the leaf. OnGuard2 also predicted with VPD unexpected alterations in K+ channel activities and changes in stomatal conductance of the slac1 Cl- channel and ost2 H+-ATPase mutants, which we verified experimentally. OnGuard2 thus bridges the micro-macro divide, offering a powerful tool with which to explore the links between guard cell homeostasis, stomatal dynamics, and foliar transpiration.
Assuntos
Arabidopsis/metabolismo , Umidade , Folhas de Planta/metabolismo , Estômatos de Plantas/metabolismo , Transdução de Sinais , Arabidopsis/citologia , Arabidopsis/genética , Transporte de Íons , Cinética , Modelos Biológicos , Mutação , Folhas de Planta/citologia , Folhas de Planta/genética , Estômatos de Plantas/genética , Transpiração Vegetal/genética , Pressão de Vapor , Água/metabolismoRESUMO
SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) proteins drive vesicle fusion and contribute to homoeostasis, pathogen defense, cell expansion, and growth in plants. In Arabidopsis (Arabidopsis thaliana), two homologous Qa-SNAREs, SYNTAXIN OF PLANTS121 (SYP121) and SYP122, facilitate the majority of secretory traffic to the plasma membrane, and the single mutants are indistinguishable from wild-type plants in the absence of stress, implying a redundancy in their functions. Nonetheless, several studies suggest differences among the secretory cargo of these SNAREs. To address this issue, we conducted an analysis of the proteins secreted by cultured wild-type, syp121, and syp122 mutant Arabidopsis seedlings. Here, we report that a number of cargo proteins were associated differentially with traffic mediated by SYP121 and SYP122. The data also indicated important overlaps between the SNAREs. Therefore, we conclude that the two Qa-SNAREs mediate distinct but complementary secretory pathways during vegetative plant growth.