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Summary: How are the transcriptional events that control form actually transduced into the shape of an organism? Analysis of plant tissue mechanical properties shows that control of the extracellular matrix is key. [ABSTRACT FROM AUTHOR]

Stomata are controllable micropores formed between two adjacent guard cells (GCs) that regulate gas flow across the plant surface. 1 Grasses, among the most successful organisms on the planet and the main food crops for humanity, have GCs flanked by specialized lateral subsidiary cells (SCs). 2,3,4 SCs improve performance by acting as a local pool of ions and metabolites to drive changes in turgor pressure within the GCs that open/close the stomatal pore. 4,5,6,7,8 The 4-celled complex also involves distinctive changes in geometry, having dumbbell-shaped GCs compared with typical kidney-shaped stomata. 2,4,9 However, the degree to which this distinctive geometry contributes to improved stomatal performance, and the underlying mechanism, remains unclear. To address this question, we created a finite element method (FEM) model of a grass stomatal complex that successfully captures experimentally observed pore opening/closure. Exploration of the model, including in silico and experimental mutant analyses, supports the importance of a reciprocal pressure system between GCs and SCs for effective stomatal function, with SCs functioning as springs to restrain lateral GC movement. Our results show that SCs are not essential but lead to a more responsive system. In addition, we show that GC wall anisotropy is not required for grass stomatal function (in contrast to kidney-shaped GCs 10) but that a relatively thick GC rod region is needed to enhance pore opening. Our results demonstrate that a specific cellular geometry and associated mechanical properties are required for the effective functioning of grass stomata. [Display omitted] • Computational modeling captures the performance of grass stomata • Specialized guard and subsidiary cell shapes set system mechanics • Normal stomatal function needs a thick guard cell rod but not wall anisotropy • Full pore closure in grasses mechanically requires subsidiary cells Grasses have a specialized form of stomata, which leads to enhanced water-use efficiency. Via computational and experimental approaches, Durney et al. show that cellular geometry sets the mechanics required for effective pore closure. The results indicate that acquisition of specific cell shape was core to the evolutionary success of our major crops. [ABSTRACT FROM AUTHOR]

Summary Stomatal opening and closure depends on changes in turgor pressure acting within guard cells to alter cell shape [ 1 ]. The extent of these shape changes is limited by the mechanical properties of the cells, which will be largely dependent on the structure of the cell walls. Although it has long been observed that guard cells are anisotropic due to differential thickening and the orientation of cellulose microfibrils [ 2 ], our understanding of the composition of the cell wall that allows them to undergo repeated swelling and deflation remains surprisingly poor. Here, we show that the walls of guard cells are rich in un-esterified pectins. We identify a pectin methylesterase gene, PME6 , which is highly expressed in guard cells and required for stomatal function. pme6-1 mutant guard cells have walls enriched in methyl-esterified pectin and show a decreased dynamic range in response to triggers of stomatal opening/closure, including elevated osmoticum, suggesting that abrogation of stomatal function reflects a mechanical change in the guard cell wall. Altered stomatal function leads to increased conductance and evaporative cooling, as well as decreased plant growth. The growth defect of the pme6-1 mutant is rescued by maintaining the plants in elevated CO 2 , substantiating gas exchange analyses, indicating that the mutant stomata can bestow an improved assimilation rate. Restoration of PME6 rescues guard cell wall pectin methyl-esterification status, stomatal function, and plant growth. Our results establish a link between gene expression in guard cells and their cell wall properties, with a corresponding effect on stomatal function and plant physiology. [ABSTRACT FROM AUTHOR]

Stomata regulate plant water use and photosynthesis by controlling leaf gas exchange. They do this by reversibly opening the pore formed by two adjacent guard cells, with the limits of this movement ultimately set by the mechanical properties of the guard cell walls and surrounding epidermis. 1,2 A body of evidence demonstrates that the methylation status and cellular patterning of pectin wall polymers play a core role in setting the guard cell mechanical properties, with disruption of the system leading to poorer stomatal performance. 3–6 Here we present genetic and biochemical data showing that wall arabinans modulate guard cell flexibility and can be used to engineer stomata with improved performance. Specifically, we show that a short-chain linear arabinan epitope associated with the presence of rhamnogalacturonan I in the guard cell wall is required for full opening of the stomatal pore. Manipulations leading to the novel accumulation of longer-chain arabinan epitopes in guard cell walls led to an increase in the maximal pore aperture. Using computational modeling combined with atomic force microscopy, we show that this phenotype reflected a decrease in wall matrix stiffness and, consequently, increased flexing of the guard cells under turgor pressure, generating larger, rounder stomatal pores. Our results provide theoretical and experimental support for the conclusion that arabinan side chains of pectin modulate guard cell wall stiffness, setting the limits for cell flexing and, consequently, pore aperture, gas exchange, and photosynthetic assimilation. [Display omitted] • Cell walls in stomata have a distinct composition of arabinans • Increasing the level of a specific arabinan makes the walls more flexible • Stomata with more flexible walls can open wider • Under high CO 2 , more flexible, wider stomata increase carbon assimilation rate The degree of stomatal opening is set by the mechanical properties of the guard cell walls. Carroll et al. show that wall flexibility is set by the arabinan composition, and that by manipulating arabinan polymers it is possible to engineer stomata with increased opening under elevated CO 2 , leading to increased carbon assimilation. [ABSTRACT FROM AUTHOR]

Summary: Stomatal pores evolved more than 410 million years ago [] and allowed vascular plants to regulate transpirational water loss during the uptake of CO2 for photosynthesis []. Here, we show that stomata on the sporophytes of the moss Physcomitrella patens [] respond to environmental signals in a similar way to those of flowering plants [] and that a homolog of a key signaling component in the vascular plant drought hormone abscisic acid (ABA) response [] is involved in stomatal control in mosses. Cross-species complementation experiments reveal that the stomatal ABA response of a flowering plant (Arabidopsis thaliana) mutant, lacking the ABA-regulatory protein kinase OPEN STOMATA 1 (OST1) [], is rescued by substitution with the moss P. patens homolog, PpOST1-1, which evolved more than 400 million years earlier. We further demonstrate through the targeted knockout of the PpOST1-1 gene in P. patens that its role in guard cell closure is conserved, with stomata of mutant mosses exhibiting a significantly attenuated ABA response. Our analyses indicate that core regulatory components involved in guard cell ABA signaling of flowering plants are operational in mosses and likely originated in the last common ancestor of these lineages more than 400 million years ago [], prior to the evolution of ferns []. [ABSTRACT FROM AUTHOR]

Summary It has long been accepted that differential radial thickening of guard cells plays an important role in the turgor-driven shape changes required for stomatal pore opening to occur [ 1–4 ]. This textbook description derives from an original interpretation of structure rather than measurement of mechanical properties. Here we show, using atomic force microscopy, that although mature guard cells display a radial gradient of stiffness, this is not present in immature guard cells, yet young stomata show a normal opening response. Finite element modeling supports the experimental observation that radial stiffening plays a very limited role in stomatal opening. In addition, our analysis reveals an unexpected stiffening of the polar regions of the stomata complexes, both in Arabidopsis and other plants, suggesting a widespread occurrence. Combined experimental data (analysis of guard cell wall epitopes and treatment of tissue with cell wall digesting enzymes, coupled with bioassay of guard cell function) plus modeling lead us to propose that polar stiffening reflects a mechanical, pectin-based pinning down of the guard cell ends, which restricts increase of stomatal complex length during opening. This is predicted to lead to an improved response sensitivity of stomatal aperture movement with respect to change of turgor pressure. Our results provide new insight into the mechanics of stomatal function, both negating an established view of the importance of radial thickening and providing evidence for a significant role for polar stiffening. Improved stomatal performance via altered cell-wall-mediated mechanics is likely to be of evolutionary and agronomic significance. [ABSTRACT FROM AUTHOR]