The dynamic multi-functionality of leaf water transport outside the xylem.

A surge of papers have reported low leaf vulnerability to xylem embolism during drought. Here, we focus on the less studied, and more sensitive, outside-xylem leaf hydraulic responses to multiple internal and external conditions. Studies of 34 species have resolved substantial vulnerability to dehydration of the outside-xylem pathways, and studies of leaf hydraulic responses to light also implicate dynamic outside-xylem responses. Detailed experiments suggest these dynamic responses arise at least in part from strong control of radial water movement across the vein bundle sheath. While leaf xylem vulnerability may influence leaf and plant survival during extreme drought, outside-xylem dynamic responses are important for the control and resilience of water transport and leaf water status for gas exchange and growth.

Coordinated decline of leaf hydraulic and stomatal conductances under drought is not linked to leaf xylem embolism for different grapevine cultivars.

Drought decreases water transport capacity of leaves and limits gas exchange, which involves reduced leaf leaf hydraulic conductance (Kleaf) in both the xylem and outside-xylem pathways. Some literature suggests that grapevines are hyper-susceptible to drought-induced xylem embolism. We combined Kleaf and gas exchange measurements, micro-computed tomography of intact leaves, and spatially explicit modeling of the outside-xylem pathways to evaluate the role of vein embolism and Kleaf in the responses of two different grapevine cultivars to drought. Cabernet Sauvignon and Chardonnay exhibited similar vulnerabilities of Kleaf and gs to dehydration, decreasing substantially prior to leaf xylem embolism. Kleaf and gs decreased by 80% for both cultivars by Ψ leaf approximately -0.7 MPa and -1.2 MPa, respectively, while leaf xylem embolism initiated around Ψ leaf = -1.25 MPa in the midribs and little to no embolism was detected in minor veins even under severe dehydration for both cultivars. Modeling results indicated that reduced membrane permeability associated with a Casparian-like band in the leaf vein bundle sheath would explain declines in Kleaf of both cultivars. We conclude that during moderate water stress, changes in the outside-xylem pathways, rather than xylem embolism, are responsible for reduced Kleaf and gs. Understanding this mechanism could help to ensure adequate carbon capture and crop performance under drought.

The Causes of Leaf Hydraulic Vulnerability and Its Influence on Gas Exchange in Arabidopsis thaliana.

The influence of the dynamics of leaf hydraulic conductance (K leaf) diurnally and during dehydration on stomatal conductance and photosynthesis remains unclear. Using the model species Arabidopsis (Arabidopsis thaliana ecotype Columbia-0), we applied a multitiered approach including physiological measurements, high-resolution x-ray microcomputed tomography, and modeling at a range of scales to characterize (1) K leaf decline during dehydration; (2) its basis in the hydraulic conductances of leaf xylem and outside-xylem pathways (K ox); (3) the dependence of its dynamics on irradiance; (4) its impact on diurnal patterns of stomatal conductance and photosynthetic rate; and (5) its influence on gas exchange and survival under simulated drought regimes. Arabidopsis leaves showed strong vulnerability to dehydration diurnally in both gas exchange and hydraulic conductance, despite lack of xylem embolism or conduit collapse above the turgor loss point, indicating a pronounced sensitivity of K ox to dehydration. K leaf increased under higher irradiance in well-hydrated leaves across the full range of water potential, but no shift in K leaf vulnerability was observed. Modeling indicated that responses to dehydration and irradiance are likely attributable to changes in membrane permeability and that a dynamic K ox would contribute strongly to stomatal closure, improving performance, survival, and efficient water use during drought. These findings for Columbia-0 provide a baseline for assessing variation across genotypes in hydraulic traits and their influence on gas exchange during dehydration.

Outside-Xylem Vulnerability, Not Xylem Embolism, Controls Leaf Hydraulic Decline during Dehydration.

Leaf hydraulic supply is crucial to maintaining open stomata for CO2 capture and plant growth. During drought-induced dehydration, the leaf hydraulic conductance (Kleaf) declines, which contributes to stomatal closure and, eventually, to leaf death. Previous studies have tended to attribute the decline of Kleaf to embolism in the leaf vein xylem. We visualized at high resolution and quantified experimentally the hydraulic vulnerability of xylem and outside-xylem pathways and modeled their respective influences on plant water transport. Evidence from all approaches indicated that the decline of Kleaf during dehydration arose first and foremost due to the vulnerability of outside-xylem tissues. In vivo x-ray microcomputed tomography of dehydrating leaves of four diverse angiosperm species showed that, at the turgor loss point, only small fractions of leaf vein xylem conduits were embolized, and substantial xylem embolism arose only under severe dehydration. Experiments on an expanded set of eight angiosperm species showed that outside-xylem hydraulic vulnerability explained 75% to 100% of Kleaf decline across the range of dehydration from mild water stress to beyond turgor loss point. Spatially explicit modeling of leaf water transport pointed to a role for reduced membrane conductivity consistent with published data for cells and tissues. Plant-scale modeling suggested that outside-xylem hydraulic vulnerability can protect the xylem from tensions that would induce embolism and disruption of water transport under mild to moderate soil and atmospheric droughts. These findings pinpoint outside-xylem tissues as a central locus for the control of leaf and plant water transport during progressive drought.

Leaf vein xylem conduit diameter influences susceptibility to embolism and hydraulic decline.

Ecosystems worldwide are facing increasingly severe and prolonged droughts during which hydraulic failure from drought-induced embolism can lead to organ or whole plant death. Understanding the determinants of xylem failure across species is especially critical in leaves, the engines of plant growth. If the vulnerability segmentation hypothesis holds within leaves, higher order veins that are most terminal in the plant hydraulic system should be more susceptible to embolism to protect the rest of the water transport system. Increased vulnerability in the higher order veins would also be consistent with these experiencing the greatest tensions in the plant xylem network. To test this hypothesis, we combined X-ray micro-computed tomography imaging, hydraulic experiments, cross-sectional anatomy and 3D physiological modelling to investigate how embolisms spread throughout petioles and vein orders during leaf dehydration in relation to conduit dimensions. Decline of leaf xylem hydraulic conductance (Kx ) during dehydration was driven by embolism initiating in petioles and midribs across all species, and Kx vulnerability was strongly correlated with petiole and midrib conduit dimensions. Our simulations showed no significant impact of conduit collapse on Kx decline. We found xylem conduit dimensions play a major role in determining the susceptibility of the leaf water transport system during strong leaf dehydration.

Grapevine petioles are more sensitive to drought induced embolism than stems: evidence from in vivo MRI and microcomputed tomography observations of hydraulic vulnerability segmentation.

The 'hydraulic vulnerability segmentation' hypothesis predicts that expendable distal organs are more susceptible to water stress-induced embolism than the main stem of the plant. In the current work, we present the first in vivo visualization of this phenomenon. In two separate experiments, using magnetic resonance imaging or synchrotron-based microcomputed tomography, grapevines (Vitis vinifera) were dehydrated while simultaneously scanning the main stems and petioles for the occurrence of emboli at different xylem pressures (Ψx ). Magnetic resonance imaging revealed that 50% of the conductive xylem area of the petioles was embolized at a Ψx of -1.54 MPa, whereas the stems did not reach similar losses until -1.9 MPa. Microcomputed tomography confirmed these findings, showing that approximately half the vessels in the petioles were embolized at a Ψx of -1.6 MPa, whereas only few were embolized in the stems. Petioles were shown to be more resistant to water stress-induced embolism than previously measured with invasive hydraulic methods. The results provide the first direct evidence for the hydraulic vulnerability segmentation hypothesis and highlight its importance in grapevine responses to severe water stress. Additionally, these data suggest that air entry through the petiole into the stem is unlikely in grapevines during drought.

Variations in xylem embolism susceptibility under drought between intact saplings of three walnut species.

A germplasm collection containing varied Juglans genotypes holds potential to improve drought resistance of plant materials for commercial production. We used X-ray computed microtomography to evaluate stem xylem embolism susceptibility/repair in relation to vessel anatomical features (size, arrangement, connectivity and pit characteristics) in 2-year-old saplings of three Juglans species. In vivo analysis revealed interspecific variations in embolism susceptibility among Juglans microcarpa, J. hindsii (both native to arid habitats) and J. ailantifolia (native to mesic habitats). Stem xylem of J. microcarpa was more resistant to drought-induced embolism as compared with J. hindsii and J. ailantifolia (differences in embolism susceptibility among older and current year xylem were not detected in any species). Variations in most vessel anatomical traits were negligible among the three species; however, we detected substantial interspecific differences in intervessel pit characteristics. As compared with J. hindsii and J. ailantifolia, low embolism susceptibility in J. microcarpa was associated with smaller pit size in larger diameter vessels, a smaller area of the shared vessel wall occupied by pits, lower pit frequency and no changes in pit characteristics as vessel diameters increased. Changes in amount of embolized vessels following 40 days of re-watering were minor in intact saplings of all three species highlighting that an embolism repair mechanism did not contribute to drought recovery. In conclusion, our data indicate that interspecific variations in drought-induced embolism susceptibility are associated with species-specific pit characteristics, and these traits may provide a future target for breeding efforts aimed at selecting walnut germplasm with improved drought resistance.