Scott A. M. McAdam

Passive Origins of Stomatal Control in Vascular Plants

The transition from passive to active metabolic control of stomata and plant water balance occurred about 360 million years ago. Carbon and water flow between plants and the atmosphere is regulated by the opening and closing of minute stomatal pores in surfaces of leaves. By changing the aperture of stomata, plants regulate water loss and photosynthetic carbon gain in response to many environmental stimuli, but stomatal movements cannot yet be reliably predicted. We found that the complexity that characterizes stomatal control in seed plants is absent in early-diverging vascular plant lineages. Lycophyte and fern stomata are shown to lack key responses to abscisic acid and epidermal cell turgor, making their behavior highly predictable. These results indicate that a fundamental transition from passive to active metabolic control of plant water balance occurred after the divergence of ferns about 360 million years ago.

Evolution of stomatal responsiveness to CO2 and optimization of water‐use efficiency among land plants

The stomata of angiosperms respond to changes in ambient atmospheric concentrations of CO(2) (C(a)) in ways that appear to optimize water-use efficiency. It is unknown where in the history of land plants this important stomatal control mechanism evolved. Here, we test the hypothesis that major clades of plants have distinct stomatal sensitivities to C(a) reflecting a relatively recent evolution of water-use optimization in derived angiosperms. Responses of stomatal conductance (g(s)) to step changes between elevated, ambient and low C(a) (600, 380 and 100 micromol mol(-1), respectively) were compared in a phylogenetically and ecologically diverse range of higher angiosperms, conifers, ferns and lycopods. All species responded to low C(a) by increasing g(s) but only angiosperm stomata demonstrated a significant closing response when C(a) was elevated to 600 micromol mol(-1). As a result, angiosperms showed significantly greater increases in water-use efficiency under elevated C(a) than the other lineages. The data suggest that the angiosperms have mechanisms for detecting and responding to increases in C(a) that are absent from earlier diverging lineages, and these mechanisms impart a greater capacity to optimize water-use efficiency.

Stomatal innovation and the rise of seed plants

Stomatal valves on the leaves of vascular plants not only prevent desiccation but also dynamically regulate water loss to maintain efficient daytime water use. This latter process involves sophisticated active control of stomatal aperture that may be absent from early-branching plant clades. To test this hypothesis, we compare the stomatal response to light intensity in 13 species of ferns and lycophytes with a diverse sample of seed plants to determine whether the capacity to optimise water use is an ancestral or derived feature of stomatal physiology. We found that in seed plants, the ratio of photosynthesis to water use remained high and constant at different light intensities, but fern and lycophyte stomata were incapable of sustaining homeostatic water use efficiency. We conclude that efficient water use in early seed plants provided them with a competitive advantage that contributed to the decline of fern and lycophyte dominated-ecosystems in the late Paleozoic.

Conifer species adapt to low-rainfall climates by following one of two divergent pathways

Significance A major determinant of plant species distribution on Earth is a specific tolerance to soil drying, yet there are currently no functional or anatomical characteristics that can predict species’ requirement for rainfall. This study examines the systems responsible for controlling water delivery and water loss in the leaves of conifers and finds functional evidence of how conifers have evolved in drying climates over the course of the last 150 million years. Two “strategies” for conserving water during water stress emerged. One group relied on the plant hormone abscisic acid to maintain stomata closed during sustained drought, and another, more derived group allowed leaves to dehydrate and resisted damage by producing a water transport system capable of functioning under the extreme tension that develops in water-stressed plants. Water stress is one of the primary selective forces in plant evolution. There are characters often cited as adaptations to water stress, but links between the function of these traits and adaptation to drying climates are tenuous. Here we combine distributional, climatic, and physiological evidence from 42 species of conifers to show that the evolution of drought resistance follows two distinct pathways, both involving the coordinated evolution of tissues regulating water supply (xylem) and water loss (stomatal pores) in leaves. Only species with very efficient stomatal closure, and hence low minimum rates of water loss, inhabit dry habitats, but species diverged in their apparent mechanism for maintaining closed stomata during drought. An ancestral mechanism found in Pinaceae and Araucariaceae species relies on high levels of the hormone abscisic acid (ABA) to close stomata during water stress. A second mechanism, found in the majority of Cupressaceae species, uses leaf desiccation rather than high ABA levels to close stomata during sustained water stress. Species in the latter group were characterized by xylem tissues with extreme resistance to embolism but low levels of foliar ABA after 30 d without water. The combination of low levels of ABA under stress with cavitation-resistant xylem enables these species to prolong stomatal opening during drought, potentially extending their photosynthetic activity between rainfall events. Our data demonstrate a surprising simplicity in the way conifers evolved to cope with water shortage, indicating a critical interaction between xylem and stomatal tissues during the process of evolution to dry climates.

Fern and Lycophyte Guard Cells Do Not Respond to Endogenous Abscisic Acid

Abscisic acid (ABA) is widely known to regulate stomatal movement. This study shows that despite the augmentation of ABA in fern and lycophyte tissues during drought, their guard cells respond passively to plant water content and not ABA levels. Stomatal insensitivity in these basal plant lineages supports the concept that the link between ABA and stomatal control is derived in seed plants. 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.

The Evolution of Mechanisms Driving the Stomatal Response to Vapor Pressure Deficit

The mechanism for a stomatal response to vapor pressure deficit evolved from a passive regulation in basal vascular plants to mediation by ABA in the earliest angiosperms. Stomatal responses to vapor pressure deficit (VPD) are a principal means by which vascular land plants regulate daytime transpiration. While much work has focused on characterizing and modeling this response, there remains no consensus as to the mechanism that drives it. Explanations range from passive regulation by leaf hydration to biochemical regulation by the phytohormone abscisic acid (ABA). We monitored ABA levels, leaf gas exchange, and water status in a diversity of vascular land plants exposed to a symmetrical, mild transition in VPD. The stomata in basal lineages of vascular plants, including gymnosperms, appeared to respond passively to changes in leaf water status induced by VPD perturbation, with minimal changes in foliar ABA levels and no hysteresis in stomatal action. In contrast, foliar ABA appeared to drive the stomatal response to VPD in our angiosperm samples. Increased foliar ABA level at high VPD in angiosperm species resulted in hysteresis in the recovery of stomatal conductance; this was most pronounced in herbaceous species. Increased levels of ABA in the leaf epidermis were found to originate from sites of synthesis in other parts of the leaf rather than from the guard cells themselves. The transition from a passive regulation to ABA regulation of the stomatal response to VPD in the earliest angiosperms is likely to have had critical implications for the ecological success of this lineage.

Abscisic acid mediates a divergence in the drought response of two conifers.

Differences in drought survival strategies of conifer species are linked to the interaction between hormone dynamics and water tension in the water transport system. During water stress, stomatal closure occurs as water tension and levels of abscisic acid (ABA) increase in the leaf, but the interaction between these two drivers of stomatal aperture is poorly understood. We investigate the dynamics of water potential, ABA, and stomatal conductance during the imposition of water stress on two drought-tolerant conifer species with contrasting stomatal behavior. Rapid rehydration of excised shoots was used as a means of differentiating the direct influences of ABA and water potential on stomatal closure. Pinus radiata (Pinaceae) was found to exhibit ABA-driven stomatal closure during water stress, resulting in strongly isohydric regulation of water loss. By contrast, stomatal closure in Callitris rhomboidea (Cupressaceae) was initiated by elevated foliar ABA, but sustained water stress saw a marked decline in ABA levels and a shift to water potential-driven stomatal closure. The transition from ABA to water potential as the primary driver of stomatal aperture allowed C. rhomboidea to rapidly recover gas exchange after water-stressed plants were rewatered, and was associated with a strongly anisohydric regulation of water loss. These two contrasting mechanisms of stomatal regulation function in combination with xylem vulnerability to produce highly divergent strategies of water management. Species-specific ABA dynamics are proposed as a central component of drought survival and ecology.

Separating active and passive influences on stomatal control of transpiration

Stomatal closure during water stress in the conifer Metasequoia glyptostroboides transitions from being entirely passive under moderate water stress to predominantly active, mediated by the level of foliar ABA, under more severe water stress. Motivated by studies suggesting that the stomata of ferns and lycophytes do not conform to the standard active abscisic acid (ABA) -mediated stomatal control model, we examined stomatal behavior in a conifer species (Metasequoia glyptostroboides) that is phylogenetically midway between the fern and angiosperm clades. Similar to ferns, daytime stomatal closure in response to moderate water stress seemed to be a passive hydraulic process in M. glyptostroboides immediately alleviated by rehydrating excised shoots. Only after prolonged exposure to more extreme water stress did active ABA-mediated stomatal closure become important, because foliar ABA production was triggered after leaf turgor loss. The influence of foliar ABA on stomatal conductance and stomatal aperture was highly predictable and additive with the passive hydraulic influence. M. glyptostroboides thus occupies a stomatal behavior type intermediate between the passively controlled ferns and the characteristic ABA-dependent stomatal closure described in angiosperm herbs. These results highlight the importance of considering phylogeny as a major determinant of stomatal behavior.

Shoot-derived abscisic acid promotes root growth.

The phytohormone abscisic acid (ABA) plays a major role in regulating root growth. Most work to date has investigated the influence of root-sourced ABA on root growth during water stress. Here, we tested whether foliage-derived ABA could be transported to the roots, and whether this foliage-derived ABA had an influence on root growth under well-watered conditions. Using both application studies of deuterium-labelled ABA and reciprocal grafting between wild-type and ABA-biosynthetic mutant plants, we show that both ABA levels in the roots and root growth in representative angiosperms are controlled by ABA synthesized in the leaves rather than sourced from the roots. Foliage-derived ABA was found to promote root growth relative to shoot growth but to inhibit the development of lateral roots. Increased root auxin (IAA) levels in plants with ABA-deficient scions suggest that foliage-derived ABA inhibits root growth through the root growth-inhibitor IAA. These results highlight the physiological and morphological importance, beyond the control of stomata, of foliage-derived ABA. The use of foliar ABA as a signal for root growth has important implications for regulating root to shoot growth under normal conditions and suggests that leaf rather than root hydration is the main signal for regulating plant responses to moisture.

Stomatal responses to vapour pressure deficit are regulated by high speed gene expression in angiosperms

Plants dynamically regulate water use by the movement of stomata on the surface of leaves. Stomatal responses to changes in vapour pressure deficit (VPD) are the principal regulator of daytime transpiration and water use efficiency in land plants. In angiosperms, stomatal responses to VPD appear to be regulated by the phytohormone abscisic acid (ABA), yet the origin of this ABA is controversial. After a 20 min exposure of plants, from three diverse angiosperm species, to a doubling in VPD, stomata closed, foliar ABA levels increased and the expression of the gene encoding the key, rate-limiting carotenoid cleavage enzyme (9-cis-epoxycarotenoid dioxygenase, NCED) in the ABA biosynthetic pathway was significantly up-regulated. The NCED gene was the only gene in the ABA biosynthetic pathway to be up-regulated over the short time scale corresponding to the response of stomata. The closure of stomata and rapid increase in foliar ABA levels could not be explained by the release of ABA from internal stores in the leaf or the hydrolysis of the conjugate ABA-glucose ester. These results implicate an extremely rapid de novo biosynthesis of ABA, mediated by a single gene, as the means by which angiosperm stomata respond to natural changes in VPD.