**1. Introduction**

It is generally accepted that plant cells expand by slow irreversible deformation of their cell walls as a result of stresses generated in the walls by the internal turgor pressure of the cell [1], and thus their growth can be framed as a biomechanical interaction between the properties of the wall of a cell and its turgor pressure. Water stress is expected to affect plant growth as a result of effects on either wall stress or wall mechanical behavior.

The turgor pressure of a cell is determined by its internal osmotic pressure and the water potential outside the cell. For a non-growing cell, it is simply the sum of these terms; however, in a growing cell there must be an imbalance favoring water movement into the cell, although the difference is likely to be slight at a cellular level. Because water potential influences turgor pressure, it is clear that the water status of a plant can alter wall stress and that this can affect the rate of plant growth; however, it should be noted that in many cases the concentration of cellular solutes increases to maintain turgor pressure [2]. There are also many examples of the active modulation of wall mechanical properties in response to signals associated with water stress or water stress itself (e.g., [3]). However, even though water makes up the majority of the primary walls of plant cells by mass (typically >80%, [4]), the direct effects of the water status of a plant on its wall properties have been relatively unexplored. Given the proportion of primary cell walls of plants that water comprises, it would be surprising if the volume that it occupies does not contribute to the mechanical behavior of the wall.

There is good reason to expect that changes in the water content of plant cell walls could affect their mechanical behavior directly. In polymer matrices, the free volume

**Citation:** Thompson, D.S.; Islam, A. Plant Cell Wall Hydration and Plant Physiology: An Exploration of the Consequences of Direct Effects of Water Deficit on the Plant Cell Wall. *Plants* **2021**, *10*, 1263. https:// doi.org/10.3390/plants10071263

Academic Editors: Penélope García-Angulo and Asier Largo-Gosens

Received: 21 May 2021 Accepted: 15 June 2021 Published: 22 June 2021

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**Copyright:** © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

between polymer molecules is thought to have substantial effects on their properties and behavior. This effect is routinely exploited to control the properties of plastics by the addition of the smaller spacing molecules, termed plasticizers (e.g., [5]). Plant cell walls comprise a composite of polymers, with the space between them generally occupied by water (although water may be displaced by other materials in secondary wall deposition).Therefore, water appears analogous to plasticizers in synthetic plastics. For example, pectin films become several-fold stiffer as their degree of hydration is reduced [6].

Tests of the effect of altering the water content on cell walls of rye coleoptiles [7] and sunflower hypocotyls [8] by using osmotica have shown the rate at which walls extended under constant load was reduced as water was removed from the walls and that these effects occurred at water potentials that plants might be expected to experience under "field" conditions (−0.4 MPa in sunflower). The tissue in both sets of experiments had been frozen and thawed, and so the effects of reduced turgor pressure and of cellular responses to the altered water potential could be excluded and the observed effects attributed solely to direct effects of hydration on the cell walls. Substantial reductions in hydration were found in tomato fruit pericarp cell wall material at even more moderate water potentials, with hydration decreasing by more than 50% as the water potential was reduced from 0 MPa to −0.15 MPa [9].

If these effects occur in vivo, changes in wall hydration, spacing and tissue biomechanics as a result of plant water status would be expected to have a direct impact on plant growth. Additionally, reducing the available space in the wall might impede the movement of enzymes and other molecules within the wall [10] and amplify the water potential gradients across tissues if apoplastic hydraulic conductivity decreases because of reduced wall area (e.g., [11]). It would therefore be surprising if responses to water availability do not include measures to maintain wall hydration or to mitigate the consequences of changes. The exploration of such adaptations may shed light on plant responses to water availability and perhaps also offer ways to maintain crop yields and improve resilience when water supplies are limited.

Such measures might include changes in wall polysaccharide composition, structure or charge to alter the relationship between wall water content and water potential, regulation of the osmotic pressure, pH or ion concentrations in the wall space to maintain hydration, or wall "loosening" to minimize any effects of hydration changes on extensibility.

One potential mechanism for controlling the water content of a cell wall is by altering its composition. In principle, this might involve changes in which polysaccharides are present or their relative proportions, but changes in polymer length and the number and composition of branches may also alter these interactions. For example, the quantity of water bound per monosaccharide unit of chitosan, alginate and cellulose in paper differs, as does the way that this relationship changes with water activity [12], such that they dehydrate at different rates as the water potential is reduced. In practice, the pectins in the wall possess the greatest chemical and structural complexity and thus potentially offer multiple mechanisms for modulating these relationships, as does the degree of esterification of pectic acidic groups.

The primary factors determining this relationship between water potential (or water activity) and the quantity of water associated with polymers are the space occupied by the polymer (itself affected by polymer length, both the number and length of branches and the mobility of polymer segments) and the strength of interactions between the polymer functional groups and water [12]. Interactions with groups at chain ends (and therefore the number of chain ends) may also be important [13]. All of these properties could be modulated by alterations in wall biosynthesis or by enzymes acting upon existing wall material.

In addition to the individual interactions between the polymers and water, the cell wall as a whole has properties resulting from interactions between its polymer components, conferring resistance to compression or conversely reducing chain separation or mobility. It certainly seems that a number of wall components have roles in maintaining the spacing

required for correct wall function [14]. This "scaffolding" may be necessary to prevent lateral collapse of walls as they deform, but also potentially to prevent compression of walls by the turgor pressures of adjacent cells pushing against one another in tissues exhibiting tissue pressure/tension [15]. Such effects on hydration as a result of interactions between polymers would be expected to include the "egg box" bridges formed by divalent cations between pectic uronic acid groups [16] as well as factors affecting the strength of these interactions, such as ionic strength and pH, if this alters the density of charged groups [9].

Cell wall enzymes offer more rapid mechanisms for modifying the relationship between water potential and wall water content than could be achieved by bulk changes in wall composition. Many types of modification could have such effects, including (but not limited to) changes in polymer molecular weight, degree of polysaccharide branching and length of branches, the number of polymer ends, and the density of charged groups. Another group of cell wall enzymes of considerable interest are the wall loosening proteins known as expansins, because it has been observed that in addition to making plant cell walls and other cellulosic materials more extensible at pH < 5.5 (e.g., [17]), expansins also cause swelling in these materials [18,19]. It therefore seems likely that, in principle, expansins could maintain or increase wall spacing under conditions of water stress.

The following experiments explore the effects of water potential on water content and mechanical behavior and of wall composition and expansins, and the interaction between them, upon these relationships in both plant materials and synthetic models of plant cell walls based on bacterial cellulose. These data establish that the water potentials that plants may experience under field conditions can directly affect the mechanical characteristics of their cell walls in ways that would be expected to affect plant growth and development in vivo and that these effects can be modulated by wall composition and expansin activity.
