Stomatal morphology and physiology explain varied sensitivity to abscisic acid across vascular plant lineages

Abscisic acid (ABA) can induce rapid stomatal closure in seed plants, but the action of this hormone on the stomata of fern and lycophyte species remains equivocal. Here, ABA-induced stomatal closure, signaling components, guard cell K⁺ and Ca²⁺ fluxes, vacuolar and actin cytoskeleton dynamics, and the permeability coefficient of guard cell protoplasts (P_f) were analyzed in species spanning the diversity of vascular land plants including 11 seed plants, 6 ferns, and 1 lycophyte. We found that all 11 seed plants exhibited ABA-induced stomatal closure, but the fern and lycophyte species did not. ABA-induced hydrogen peroxide elevation was observed in all species, but the signaling pathway downstream of nitric oxide production, including ion channel activation, was only observed in seed plants. In the angiosperm faba bean (Vicia faba), ABA application caused large vacuolar compartments to disaggregate, actin filaments to disintegrate into short fragments and P_f to increase. None of these changes was observed in the guard cells of the fern Matteuccia struthiopteris and lycophyte Selaginella moellendorffii treated with ABA, but a hypertonic osmotic solution did induce stomatal closure in fern and the lycophyte. Our results suggest that there is a major difference in the regulation of stomata between the fern and lycophyte plants and the seed plants. Importantly, these findings have uncovered the physiological and biophysical mechanisms that may have been responsible for the evolution of a stomatal response to ABA in the earliest seed plants.

Physiological and biophysical evidence for insensitivity of stomata to abscisic acid in ferns and lycophytes supports stomatal responsiveness to abscisic acid evolved after the divergence of ferns.     

Fern and lycophyte guard cells do not respond to endogenous abscisic acid

Stomatal guard cells regulate plant photosynthesis and transpiration. Central to the control of seed plant stomatal movement is the phytohormone abscisic acid (ABA); however, differences in the sensitivity of guard cells to this ubiquitous chemical have been reported across land plant lineages. Using a phylogenetic approach to investigate guard cell control, we examined the diversity of stomatal responses to endogenous ABA and leaf water potential during water stress. We show that although all species respond similarly to leaf water deficit in terms of enhanced levels of ABA and closed stomata, the function of fern and lycophyte stomata diverged strongly from seed plant species upon rehydration. When instantaneously rehydrated from a water-stressed state, fern and lycophyte stomata rapidly reopened to predrought levels despite the high levels of endogenous ABA in the leaf. In seed plants under the same conditions, high levels of ABA in the leaf prevented rapid reopening of stomata. We conclude that endogenous ABA synthesized by ferns and lycophytes plays little role in the regulation of transpiration, with stomata passively responsive to leaf water potential. These results support a gradualistic model of stomatal control evolution, offering opportunities for molecular and guard cell biochemical studies to gain further insights into stomatal control.     

西双版纳热带雨林中两种生态型蕨类植物的光合特性比较研究

通过比较分布于西双版纳热带雨林林下生境中的附生鸟巢蕨(Neottopteris nidus)和地生网脉铁角蕨(Asplenium finlaysonianum)的光合特征和光合诱导特性, 来研究不同生态型蕨类植物的光斑利用策略。研究结果表明, 2种蕨类植物的最大净光合速率、暗呼吸速率、表观量子效率、光饱和点和光补偿点没有显著差异, 但网脉铁角蕨的最大气孔导度远远高于鸟巢蕨, 表明后者具有更强的光合水分利用效率。在暗处理3小时接着光照(光强为20 mmol .m-2.s-1)30分钟后, 网脉铁角蕨的初始气孔导度显著高于鸟巢蕨。连续照射饱和强光后, 网脉铁角蕨达到最大净光合速率50%(T50%)和90%的时间(T90%)比鸟巢蕨短: 网脉铁角蕨和鸟巢蕨的T50%分别为0.57和5.31分钟, T90%分别为5.85和26.33分钟。诱导过程中, 气孔导度对强光的响应明显滞后于净光合速率。鸟巢蕨达到最大气孔导度的时间明显比网脉铁角蕨慢, 但在光合诱导消失过程中2种蕨类植物的 光合诱导维持能力却没有显著差异。上述结果表明, 与大多数地生林下植物(如网脉铁角蕨)相比, 附生鸟巢蕨的水分保护比碳获得更重要, 但却限制了附生蕨对光斑的利用。     

西双版纳热带雨林中两种生态型蕨类植物的光合特性比较研究

通过比较分布于西双版纳热带雨林林下生境中的附生鸟巢蕨（Neottopteris nidus）和地生网脉铁角蕨（Asplenium finlaysonianum）的光合特征和光合诱导特性,来研究不同生态型蕨类植物的光斑利用策略。研究结果表明,2种蕨类植物的最大净光合速率、暗呼吸速率、表观量子效率、光饱和点和光补偿点没有显著差异,但网脉铁角蕨的最大气孔导度远远高于鸟巢蕨,表明后者具有更强的光合水分利用效率。在暗处理3／J＇,时接着光照（光强为20I~mol-m-2,s。‘）30分钟后,网脉铁角蕨的初始气孔导度显著高于鸟巢蕨。连续照射饱和强光后,网脉铁角蕨达到最大净光合速率50％（T50％）和90％的时间（T90%）比鸟巢蕨短：网脉铁角蕨和鸟巢蕨的T50%分别为0．57和5．31分钟,T90％分别为5．85和26．33分钟。诱导过程中,气孔导度对强光的响应明显滞后于净光合速率。鸟巢蕨达到最大气孔导度的时间明显比网脉铁角蕨慢,但在光合诱导消失过程中2种蕨类植物的光合诱导维持能力却没有显著差异。上述结果表明,与大多数地生林下植物（如网脉铁角蕨）相比,附生鸟巢蕨的水分保护比碳获得更重要,但却限制了附生蕨对光斑的利用。     

Hormonal dynamics contributes to divergence in seasonal stomatal behaviour in a monsoonal plant community

The plant hormone abscisic acid (ABA) is a primary regulator of plant transpiration, but its influence in determining seasonal stomatal behaviour in natural plant communities is poorly understood. We examined distantly related vascular plants growing together in a seasonally dry, monsoonal environment to determine whether ABA dynamics contributed to contrasting water use patterns in this natural setting. Regular sampling of angiosperm, cycad, conifer and fern species revealed characteristic seasonal patterns in ABA production, but these were highly distinct among species. Although no general relationship was observed between ABA levels, plant hydration or stomatal conductance among species, the seasonal dynamics in stomatal behaviour within species were predictable functions of either ABA or leaf water potential. Strong divergence in the seasonal role of ABA among species suggests that modification in ABA–stomatal interactions represents an important evolutionary pathway for adaptation in plant water use.     

弱光下生长的葡萄叶片蒸腾速率和气孔结构的变化

 植物能够对生长环境产生生态适应性,这种适应性可从气孔导度、光合速率、水分利用效率等生态指标上反映出来。为了研究葡萄蒸腾特性对弱光环境的适应性变化,本试验以‘京玉’葡萄幼苗（Vitis vinefera cv. Jingyu）为试验材料,通过遮光处理（2个处理,分别遮光65%和85%）营造弱光环境,测定了在弱光环境下生长的葡萄叶片蒸腾速率、气孔导度、水分利用效率对光照强度的响应,同时用扫描电镜技术观察了气孔的发育。结果表明,弱光环境下生长的葡萄幼苗,叶片的水势较高,但水分利用效率较低,叶片蒸腾速率和气孔导度变化对光照强度的响应缓慢,而自然光下生长的葡萄叶片则反应较迅速。通过对气孔结构的研究发现,与自然光照环境下生长的植株相比,在弱光环境下生长的葡萄幼苗,叶片下表皮的气孔横轴变宽,大小气孔之间差异减少,气孔外突,表皮细胞变大甚至扭曲,角质层变薄。说明葡萄幼苗能够对弱光环境产生适应性变化,其蒸腾特性的变化与其气孔结构的变化相关,具有一致性。     

The fern Adiantum capillus-veneris lacks stomatal responses to blue light

We investigated the responses of stomata to light in the fern Adiantum capillus-veneris, a typical species of Leptosporangiopsida. Stomata in the intact leaves of the sporophytes opened in response to red light, but they did not open when blue light was superimposed on the red light. The results were confirmed in the isolated Adiantum epidermis. The red light-induced stomatal response was not affected by the mutation of phy3, a chimeric protein of phytochrome and phototropin in this fern. The lack of a blue light-specific stomatal response was observed in three other fern species of Leptosporangiopsida, i.e. Pteris cretica, Asplenium scolopendrium and Nephrolepis auriculata. Fusicoccin, an activator of the plasma membrane H(+)-ATPase, induced both stomatal opening and H(+) release in the Adiantum epidermis. Adiantum phototropin genes AcPHOT1 and AcPHOT2 were expressed in the fern guard cells. The transformation of an Arabidopsis phot1 phot2 double mutant, which lost blue light-specific stomatal opening, with AcPHOT1 restored the stomatal response to blue light. Taken together, these results suggest that ferns of Leptosporangiopsida lack a blue light-specific stomatal response, although the functional phototropin and plasma membrane H(+)-ATPase are present in this species.     

Shade does not ameliorate drought effects on the tree fern species Dicksonia antarctica and Cyathea australis

We examined the responses of two tree fern species (Dicksonia antarctica and Cyathea australis) growing under moderate and high light regimes to short-term water deficit followed by rewatering. Under adequate water supply, morphological and photosynthetic characteristics differed between species. D. antarctica, although putatively the more shade and less drought adapted species, had greater chlorophyll a/b ratio, and greater water use efficiency and less negative δ¹³C. Both species were susceptible to water deficit regardless of the light regime showing significant decreases in photosynthetic parameters (A _max, V _cmax, J _max) and stomatal conductance (g _s ) in conjunction with decreased relative frond water content (RWC) and predawn frond water potential (Ψ_predawn). During the water deficit period, decreases in g _s in both species started one day later, and were at lower soil water content, under moderate light compared with high light. D. antarctica under moderate light was more vulnerable to drought than all other plants as was indicated by greater decreases in Ψ_predawn, lowest stomatal conductance, and photosynthetic rates. Both tree fern species were able to recover after a short but severe water stress.     

石松类和蕨类植物质体基因组结构演化研究进展

近年来, 随着测序技术的发展, 石松类和蕨类植物的核基因组、质体基因组以及线粒体基因组研究发展迅速, 质体基因组研究工作更是呈爆发式增长。截至2019年3月1日, GenBank公布的石松类和蕨类植物的175个质体基因组中, 约3/4为最近两年新增。研究内容从早期对个别质体基因组结构和序列特征的简单报道, 逐渐发展到综合性的比较基因组学和系统发育基因组学研究。目前已发表的质体基因组覆盖了石松类和蕨类植物的所有目和大部分科, 这两大类群的质体基因组结构变异和系统发育的基本框架已逐渐清晰。这些研究为我们理解维管植物的早期演化提供了重要参考。本文对石松类和蕨类植物的质体基因组结构特征进行了系统梳理, 发现其结构变异主要包括大片段倒位、IR区边界变动、基因或内含子丢失等, 其中一些结构变异可作为较高分类阶元的共衍征。RNA编辑和长片段非编码序列插入普遍存在于石松类和蕨类植物的质体基因组中, 但其起源、演化机制和功能等仍不清楚。我们对质体基因组的应用、系统发育研究中质体和核基因组的优劣性, 以及系统发育基因组学的前景进行了评述。     

Physiological framework for adaptation of stomata to CO2 from glacial to future concentrations

In response to short-term fluctuations in atmospheric CO(2) concentration, c(a), plants adjust leaf diffusive conductance to CO(2), g(c), via feedback regulation of stomatal aperture as part of a mechanism for optimizing CO(2) uptake with respect to water loss. The operational range of this elaborate control mechanism is determined by the maximum diffusive conductance to CO(2), g(c(max)), which is set by the size (S) and density (number per unit area, D) of stomata on the leaf surface. Here, we show that, in response to long-term exposure to elevated or subambient c(a), plants alter g(c(max)) in the direction of the short-term feedback response of g(c) to c(a) via adjustment of S and D. This adaptive feedback response to c(a), consistent with long-term optimization of leaf gas exchange, was observed in four species spanning a diverse taxonomic range (the lycophyte Selaginella uncinata, the fern Osmunda regalis and the angiosperms Commelina communis and Vicia faba). Furthermore, using direct observation as well as flow cytometry, we observed correlated increases in S, guard cell nucleus size and average apparent 1C DNA amount in epidermal cell nuclei with increasing c(a), suggesting that stomatal and leaf adaptation to c(a) is linked to genome scaling.     

Distinct developmental mechanisms reflect the independent origins of leaves in vascular plants

Vascular plants diverged more than 400 million years ago into two lineages, the lycophytes and the euphyllophytes . Leaf-like organs evolved independently in these two groups . Microphylls in lycophytes are hypothesized to have originated as lateral outgrowths of tissue that later became vascularized (the enation theory) or through the sterilization of sporangia (the sterilization hypothesis) . Megaphylls in euphyllophytes are thought to represent modified lateral branches . The fossil record also indicates that the seed plant megaphyll evolved uniquely in the ancestor of seed plants, independent of megaphylls in ferns, because seed plants evolved from leafless progymnosperm ancestors . Surprisingly, a recent study of KNOX and ARP gene expression in a lycophyte was reported to indicate recruitment of a similar mechanism for determinacy in both types of leaves . We examined the expression of Class III HD-Zip genes in the lycophyte Selaginella kraussiana and in two gymnosperms, Ginkgo and Pseudotsuga. Our data indicate that mechanisms promoting leaf initiation, vascularization, and polarity are quite different in lycophytes and seed plants, consistent with the hypotheses that megaphylls originated as lateral branches whereas microphylls originated as tissue outgrowths.     

Light-Induced Stomatal Opening Is Affected by the Guard Cell Protein Kinase APK1b

Guard cells allow land plants to survive under restricted or fluctuating water availability. They control the exchange of gases between the external environment and the interior of the plant by regulating the aperture of stomatal pores in response to environmental stimuli such as light intensity, and are important regulators of plant productivity. Their turgor driven movements are under the control of a signalling network that is not yet fully characterised. A reporter gene fusion confirmed that the Arabidopsis APK1b protein kinase gene is predominantly expressed in guard cells. Infrared gas analysis and stomatal aperture measurements indicated that plants lacking APK1b are impaired in their ability to open their stomata on exposure to light, but retain the ability to adjust their stomatal apertures in response to darkness, abscisic acid or lack of carbon dioxide. Stomatal opening was not specifically impaired in response to either red or blue light as both of these stimuli caused some increase in stomatal conductance. Consistent with the reduction in maximum stomatal conductance, the relative water content of plants lacking APK1b was significantly increased under both well-watered and drought conditions. We conclude that APK1b is required for full stomatal opening in the light but is not required for stomatal closure.     

Complex Effects of Prescribed Fire on Restoring the Soil Water Content in a High-Elevation Riparian Meadow, Arizona

One of the largest and rarest Bebb willow (Salix bebbiana) communities in the United States occurs at Hart Prairie, Arizona. Low recruitment of the willow over the past several decades has been linked to inadequate soil water content for seed germination and seedling establishment. We tested a hypothesis that a prescribed burn would reduce biomass of and evapotranspiration by herbaceous plants, thereby increasing soil water content. Three treatments (unburned control, early‐growing season burned, late‐growing season burned) were applied in year 2001 to replicated plots in fern‐ and grass‐dominated herbaceous communities. Soil water content (0–30 cm) was measured weekly in plots during the 2001, 2002, and 2003 growing seasons. Both early‐ and late‐season burning reduced herbaceous biomass in the fern‐dominated community in 2002 and 2003 and reduced biomass in the grass‐dominated community in 2002 but not in 2003. Soil water content increased for approximately four weeks in 2001 following the early‐season burn, but the early‐season and late‐season burns reduced soil water content in both communities over much of the 2002 and 2003 growing seasons. Thus, early‐season burning may benefit willow seed germination by increasing soil water content immediately following burning but be detrimental to germination in the second and third growing seasons after burning because of drier soil. Large temporal variation in the effect of prescribed burning on soil water content will complicate the use of fire as a restoration tool to manage soil water available for threatened plants such as Bebb willow and for recharge of groundwater.     

The evolution of chloroplast genes and genomes in ferns

Most of the publicly available data on chloroplast (plastid) genes and genomes come from seed plants, with relatively little information from their sister group, the ferns. Here we describe several broad evolutionary patterns and processes in fern plastid genomes (plastomes), and we include some new plastome sequence data. We review what we know about the evolutionary history of plastome structure across the fern phylogeny and we compare plastome organization and patterns of evolution in ferns to those in seed plants. A large clade of ferns is characterized by a plastome that has been reorganized with respect to the ancestral gene order (a similar order that is ancestral in seed plants). We review the sequence of inversions that gave rise to this organization. We also explore global nucleotide substitution patterns in ferns versus those found in seed plants across plastid genes, and we review the high levels of RNA editing observed in fern plastomes.     

The stomata of the fern Adiantum capillus-veneris do not respond to CO2 in the dark and open by photosynthesis in guard cells

The stomata of the fern Adiantum capillus-veneris lack a blue light-specific opening response but open in response to red light. We investigated this light response of Adiantum stomata and found that the light wavelength dependence of stomatal opening matched that of photosynthesis. The simultaneous application of red (2 micromol m(-2) s(-1)) and far-red (50 micromol m(-2) s(-1)) light synergistically induced stomatal opening, but application of only one of these wavelengths was ineffective. Adiantum stomata did not respond to CO2 in the dark; the stomata neither opened under a low intercellular CO2 concentration nor closed under high intercellular CO2 concentration. Stomata in Arabidopsis (Arabidopsis thaliana), which were used as a control, showed clear sensitivity to CO2. In Adiantum, stomatal conductance showed much higher light sensitivity when the light was applied to the lower leaf surface, where stomata exist, than when it was applied to the upper surface. This suggests that guard cells likely sensed the light required for stomatal opening. In the epidermal fragments, red light induced both stomatal opening and K+ accumulation in guard cells, and both of these responses were inhibited by a photosynthetic inhibitor, 3-(3,4-dichlorophenyl)-1,1-dimethylurea. The stomatal opening was completely inhibited by CsCl, a K+ channel blocker. In intact fern leaves, red light-induced stomatal opening was also suppressed by 3-(3,4-dichlorophenyl)-1,1-dimethylurea. These results indicate that Adiantum stomata lack sensitivity to CO2 in the dark and that stomatal opening is driven by photosynthetic electron transport in guard cell chloroplasts, probably via K+ uptake.     

Stomatal Opening Mechanism of CAM Plants

Stomata usually open when leaves are transferred from darkness to light. However, reverse-phase stomatal opening in succulent plants has been known. CAM plants such as cacti and Opuntia ficus–indica achieve their high water use efficiency by opening their stomata during the cool, desert nights and closing them during the hot, dry days. Signal transduction pathway for stomatal opening by blue light photoreceptors including phototropins and the carotenoid pigment zeaxanthin has been suggested. Blue light regulated signal transduction pathway on stomatal opening could not be applied to CAM plants, but the most possible theory for a nocturnal response of stomata in CAM plants is photoperiodic circadian rhythm.     

The stomatal response to evaporative demand persists at night in Ricinus communis plants with high nocturnal conductance

Evidence is building that stomatal conductance to water vapour (g(s)) can be quite high in the dark in some species. However, it is unclear whether nocturnal opening reflects a mechanistic limitation (i.e. an inability to close at night) or an adaptive response (i.e. promoting water loss for reasons unrelated to carbon gain). Further, it is unclear if stomatal responses to leaf-air vapour pressure difference (D) persist in the dark. We investigated nocturnal stomatal behaviour in castor bean (Ricinus communis L.) by measuring gas exchange and stomatal responses to D in the light and in the dark. Results were compared among eight growth environments [two levels for each of three treatment variables: air saturation deficit (D(a)), light and water availability]. In most plants, stomata remained open and sensitive to D at night. g(s) was typically lower at night than in the day, whereas leaf osmotic pressure (Pi) was higher at night. In well-watered plants grown at low D(a), stomata were less sensitive to D in the dark than in the light, but the reverse was found for plants grown at high D(a). Stomata of droughted plants were less sensitive to D in the dark than in the light regardless of growth D(a). Drought also reduced g(s) and elevated Pi in both the light and the dark, but had variable effects on stomatal sensitivity to D. These results are interpreted with the aid of models of stomatal conductance.     

A survey of the mycorrhization of Southeast Asian ferns and lycophytes

The colonisation of land by plants may not have been possible without mycorrhizae, which supply the majority of land plants with nutrients, water and other benefits. In this sense, the mycorrhization of basal groups of land plants such as ferns and lycophytes is of particular interest, yet only about 9% of fern and lycophyte species have been sampled for their mycorrhization status, and no community‐level analyses exist for tropical fern communities. In the present study, we screened 170 specimens of ferns and lycophytes from Malaysia and Sulawesi (Indonesia), representing 126 species, and report the mycorrhization status for 109 species and 19 genera for the first time. Mycorrhizal colonisations were detected in 96 (56.5%) of the specimens, 85 of which corresponded to arbuscular mycorrhizae (AMF), three to dark‐septate endophytes (DSE) and four to mixed colonisations (AMF + DSE). DSE colonisations were lower than in comparable samples of ferns from the Andes, suggesting a geographical or taxonomic pattern in this type of colonisation. Epiphytes had significantly lower levels of colonisation (26.1%) than terrestrial plants (70.7%), probably due to the difficulty of establishment of mycorrhizal fungi in the canopy habitat.