The Plant Cell, January 2014, www.plantcell.org ã 2014 American Society of Plant Biologists. All rights reserved.

TEACHING TOOLS IN PLANT BIOLOGY™: LECTURE NOTES

Water Relations 1: Uptake and Transport

This is the first part of two Teaching Tools in Plant Biology to published in 1727 in the book Vegetable Staticks by the great
examine plant–water relations. Part 1 examines how plants take up plant physiologist Stephen Hales. Hales showed that the liquid
water from their environments and transport it through their bodies. transpiring from leaves is pure water, and by identifying a way to
Part 2 looks at plant responses to drought and adaptations to dry measure transpiration, he ruled out circulation as a significant
environments as well as strategies to produce sufficient food, feed, form of water movement in plants. He also demonstrated that the
and fiber within the constraints of water limitation. upward flow of water in a stem requires evaporation from leaves
and that, under most conditions, pressure from the roots is not
INTRODUCTION involved. Eduard Strasburger later demonstrated that living cells
are not required for water conduction (although, as we will see
Plants, like all living things, are mostly water. Water is the matrix later, living cells are nevertheless critical for optimizing and regulating
of life, and its availability largely determines the distribution and water conduction).
productivity of plants on earth. To survive in the dry terrestrial In the 1890s, several scientists, notably Böhm, Dixon, and Joly,
environment, plants have evolved two different strategies to converged on the idea that the driving force for the movement of
manage water uptake and transport. These are best considered water was the evaporation of water at the leaf surface, transduced
as opposite ends of a continuum, where individual plant species through the water column because of water’s chemical cohesion.
may display combinations of both, in varying degrees and at Dixon and Joly explained, “the all-sufficient cause of the elevation of
various times. the sap [is].by exerting a simple tensile stress on the liquid in the
The first is to manage water relations minimally and to vary conduits.” This idea, which came to be known as the cohesion–
cellular water content with environmental water availability. This tension theory, elegantly describes our current understanding of the
strategy is called poikilohydry; poikilo means “variable.” (The ascent of water. Dixon and Joly showed that the force exerted by
prefix is also used in poikilothermy, the strategy of varying body the evaporative pull was sufficient to raise water many meters, and
temperature with the external environment that is used by reptiles later, a similar strategy was used to measure tension in the xylem.
and other “cold-blooded” animals.) Poikilohydrous plants are able By the end of the 19th century, the idea that water moves passively
to survive periods of extensive desiccation. Most bryophytes under tension had been proposed and generally accepted, although
(mosses, liverworts, and hornworts) use this strategy, but it is also concerns about this model’s ability to explain the movement of
used by a few vascular plants, including ferns and monocots. The water up very tall trees and vines persisted. Resolving this and other
cellular mechanisms of desiccation tolerance are fascinating and questions required the development of additional methods for
covered in detail in Part 2 of this Teaching Tool. measuring xylem tension, and discussions and disagreements
The second strategy is to manage water uptake and transport, about how to interpret these various approaches persist.
in order to maintain relatively constant internal water content. This
strategy is called homoiohydry; homoio means “same.” Homo-
iohydrous plants, primarily tracheophytes, have evolved complex WATER UPTAKE AND TRANSPORT ARE GOVERNED BY
mechanisms by which to absorb, transport, and retain water; PHYSICAL LAWS
these include roots, xylem, and the water-conducting tracheary Water flows into and through plants as a continuous column of
elements, and regulated pores (stomata). To understand how water. Ultimately, the energy required to move this column of water
water moves in vascular plants, it is important to first understand comes from sunlight, which drives the evaporation of water at the
the properties that make water indispensable for life. leaf surface. As water evaporates from the top of the water column,
it exerts tension on the column that draws more water in at the
A BRIEF HISTORY OF THE STUDY OF bottom of the column. In fact, the movement of water in a plant can
PLANT–WATER RELATIONS be replicated by a physical nonliving object that has an evaporative
surface on top of a water-filled tube. The basis for this simple
In the 1620s, William Harvey demonstrated that blood in animals
mechanism, which requires no input of energy from the plant, rests
is circulated by the action of the pumping heart. In the late 17th
on the special properties of water, particularly its cohesiveness.
century, both Marcello Malpighi and Nehemiah Grew recognized
that plant vascular tissues might be conducting tissues, although
they assumed that these conduits carried air, so they became Special Properties of Water
known as tracheids or tracheary elements. The most informative
Water is cohesive, a bit like Lego. Lego bricks attach to each
early studies of the mechanism of water movement in plants were
other, so the structures you build with them are durable and stay
www.plantcell.org/cgi/doi/10.1105/tpc.114.tt0114 together; imagine trying to build similarly elaborate structures with
2 The Plant Cell

pebbles or playing cards that lack Lego’s cohesive properties. osmotic potential to negative values. The more dissolved solutes,
Water molecules stick to other water molecules because each the more negative the osmotic potential and the greater the
molecule has a partial charge separation, so that the oxygen atom tendency for water to move into the solution. Thus, one way to
at the center of the molecule has a slight negative charge and move water is to add solutes to the place you want the water to go;
flanking hydrogens have a slight positive charge; it is a polar the lower osmotic potential will draw the water toward it. A simple
molecule, but with no net charge. Due to the charge separation, way to remember this is through the expression “water follows
water molecules form intermolecular hydrogen bonds that hold salt.”
them together. The cohesiveness of water means that, compared
with other similarly sized molecules, more energy is needed to
Pressure Potential
convert water from a liquid to a gas. Thus, water is a liquid at
standard earth temperatures, whereas many other compounds of Another thermodynamic property of water is its pressure, relative
similar molecular weight are gases. Water’s polar nature also to atmospheric pressure at the earth’s surface. Because it is
means that it is a very effective solvent for highly polar molecules defined relative to atmospheric pressure, the pressure potential of
such as mineral ions as well for somewhat less polar organic the water in a container open to the atmosphere is equal to 0 MPa.
peptides and carbohydrates. On earth, if you want a molecule that If that water were put into a pressurized container, its pressure
supports life, there’s no substitute for water. potential would be a positive value, and if it were put into
a vacuum, the pressure would be a negative value relative to
atmospheric pressure. Water flows from high pressure to low
Factors That Govern the Movement of Water pressure. This can be illustrated by the use of a syringe. When
the opening of the syringe is placed into a pool of water
Diffusion and the plunger is pulled up, the pressure inside the barrel
All matter is in continuous motion. If an element is present in a is lowered and water is pulled into it. When the plunger is
higher concentration in one region, more molecules move away pushed, the pressure inside the barrel is elevated and water is
from that region than toward it. As a consequence, without any pushed out.
input of external energy, gaseous and liquid matter tends to Most plant cells are under a positive pressure, on the order of
become uniformly distributed; this phenomenon can be de- 0.5 to 1.5 MPa. This is more than the pressure of a car tire (which
scribed by Fick’s law of diffusion: is ;0.2 MPa) and manyfold higher than that experienced by
animal cells; human arterial blood pressure, for example, is less
dm dc than 0.02 MPa. This positive pressure, often called turgor pressure,
¼ 2D A
dx dx is essential for the support of many tissues. A plant wilts when it
cannot absorb enough water to maintain turgor pressure. Positive
Fick’s law states that rate of movement of a substance (dm/dx) is pressure is also essential for cell growth; for example, the
determined by its concentration gradient (dc/dx), the area through elongation of a root tip into compacted soil is driven by pressure
which it is diffusing (A), and the properties of the substance and within the cells. Changes in pressure can also affect cell movement,
the matrix through which it is diffusing (defined as the diffusion for example, by forcing open, and then releasing, the hinge of
coefficient D). The negative sign before D indicates simply that the a Venus flytrap (Dionaea muscipula) and the motor cells in the
direction of movement is away from the area of higher concentration. leaves of the sensitive plant (Mimosa pudica). By contrast, and as
described later, the water inside the conducting tissues of the
Osmosis and Osmotic Potential xylem is normally under a negative pressure (tension).

Osmosis is a special type of diffusion that applies only to water

moving across a semipermeable barrier, such as the plasma Water Potential
membrane of a cell. If we imagine a space that contains water
Water potential provides a convenient quantification of the energy
with dissolved solutes surrounded by a membrane that is
state of water, differences in which can drive water movement,
permeable to water but not solutes, then the water inside the
and it is the sum of osmotic, pressure and gravitational potentials.
membrane will essentially be at a lower concentration than that
The standard notation to express this is the water potential equation:
outside, so water will enter by diffusion. The force that drives
water inward is determined by the concentration of solutes. In cw ¼ cp þ cp þ cg
a constrained volume, the resulting pressure is the osmotic
pressure, which is defined as force (in Newtons [N]) per area
(m22). Force per area (N m22) is also a measure of pressure, for where cw indicates water potential, cπ indicates osmotic
which the standard unit is Pascals (Pa). For use in plant physiology, potential, cp indicates pressure potential, and cg indicates the
the megapascal (MPa, equivalent to 106 N m22) is a more useful gravitational potential (c is the Greek letter psi, pronounced
unit scale. Other units can be used for pressure; 1 MPa is equivalent “sigh”). The water potential of pure water at standard atmospheric
to 10 bars or 9.87 atmospheres. pressure is 0 MPa, because both the osmotic and pressure
Osmotic potential is the inverse of the osmotic pressure and is potentials are zero, by definition. The water potential can be
a property of a solution. The osmotic potential of pure water is lowered by lowering the osmotic potential (by adding solutes) or
defined as 0 MPa, and the addition of any solute lowers the the pressure potential (by applying a vacuum or other tensile
 January 2014 3

stress). The gravitational potential becomes more negative by I ¼ V=R

0.1 MPa for each 10 m of elevation; this component is often
ignored, except in the case of tall trees. The rule governing the Although this equation describes the flow of electrons (current)
movement of water across membranes is that water moves from through an electrical circuit, it also analogously describes the flow
higher to lower water potential. When no membranes are traversed, of water. Flow is determined by the difference in water potential
for example, in the column of water in the xylem or phloem, water’s between the beginning and end of the path (equivalent to the
movement is driven by the pressure gradient. The movement of voltage) divided by the resistance to the flow. An enormous
water in a transpiring plant involves the continual movement of pressure difference can only produce a small flow if the resistance
water from the soil, into the plant tissues, and ultimately out into to its flow is very large; think of a giant bucket with a pinprick-sized
the atmosphere down a continually decreasing water potential hole at the bottom. A much larger hole would give less resistance
or pressure potential gradient. and increase the rate of the flow.
Water potential values of plant tissues vary, but as typical When looking at a complex system such as a plant, it is worth
examples, the water potential of moist soil is about 20.05 MPa, that considering how resistance is affected in various segments sepa-
of a living cell in the root or leaf tissue is about 20.7 MPa, that of the rately. For example, the resistances of the root system, stem system,
substomatal cavity is about 27 MPa, and that of the atmospheric and leaf can be measured independently. The resistance of the
air varies with humidity and temperature, from (at 20°C) 215 MPa entire system is the sum of the resistances of each of the
at 90% relative humidity to 295 MPa at 50% relative humidity and segments. Thus, a system in which one component has a very
correspondingly lower with drier air. large resistance will tend to be dominated by that resistance.
Another way to think about water flow is to consider the
conductance of the system; conductance is the reciprocal of
Water Moves in Plants by Diffusion and by Bulk Flow
resistance (1/R). Although we can measure the conductance of
The movement of water by osmosis and diffusion occurs at the each part of the pathway separately, conductances cannot simply
molecular level and does not require any input of energy. However, be summed to determine the conductance of the system. Both
the movement of water by diffusion is a relatively slow process. For resistance and conductance are routinely used in plant physiology.
example, a water molecule can diffuse across a 50 μm cell in 0.6 s,
but it would take 8 years to diffuse 1 m. Thus, diffusion alone is A Note about Nomenclature
insufficient to support the needs of the typical plant for water.
Instead, vascular plants rely on bulk flow of water. Bulk flow is The general transport equation, applicable to the transport of
a much faster form of water movement, but it also requires an water, heat, and electric charge, is that flux = driving force ×
energy source to generate the required pressure gradient. proportionality constant. Fick’s law, Poiseuille’s law, and Ohm’s
Examples of the bulk flow of water are the movement of water law are all variants of the transport equation (see Davies, 1986).
in a stream (with gravity providing the energy) or out of a water Different units can be used for flux (or flow rate) as well as the
pipe (with energy provided by a pump) or the movement of proportionality constant. One commonly used proportionality
blood in the human body (with energy provided by a pump, the constant is the absolute hydraulic conductance (K), which is the
heart). ratio of the water flow rate through a plant or plant segment to
The bulk flow of water through a tube is described by one form the pressure driving the flow.
of Poiseuille’s law, which states that flow is equal to the Conductance can be restricted to a portion of the plant; for
hydraulic pressure gradient (ΔP) times π, the number of tubes example, KS indicates the conductance of the shoot and KL the
(N), and the radius of the tubes to the fourth power (r4), divided conductance of the leaf. The conductance of the root is sometimes
by 8 times the viscosity of water (η), with units in terms of volume indicated as KR and also sometimes as Lo. Conductance is also
flow per unit of time (e.g., m3 s21): sometimes abbreviated g, which is commonly used when de-
scribing the movement of CO2 and water vapor through the leaf, for
πNr4 example, stomatal conductance (gs), mesophyll cell conductance
Flow ¼ ΔP
8η (gm), and conductance of the intercellular air space (gias). In each
case, it can be helpful to remember that conductance is the
An important implication of this equation for plant physiol- reciprocal of resistance, so a higher conductance means a greater
ogists derives from the fact that flow is proportional to the flow rate per unit of driving force. A related concept is the hydraulic
fourth power of the tube radius, so a small increase in tube conductivity (e.g., ks [xylem-specific conductivity] or Lp [root
diameter leads to a tremendous increase in flow. We’ll return hydraulic conductivity]), which is calculated by dividing the flow
to this relationship when we look at the movement of water in rate by the pressure gradient over a segment of length or area
the conduits of the xylem. across which flow is occurring (see Becker et al., 1999).

Ohm’s Law Can Model the Flow of Water through a Plant AQUAPORINS, ESSENTIAL REGULATORS
OF WATER MOVEMENT
Another way to think about the flow of water in a plant is through
Ohm’s law. Ohm’s law states that current (I) equals voltage (V) For many years, the movement of water across cell membranes
divided by resistance (R). was hypothesized to involve some sort of water channel, but it
4 The Plant Cell

was not until the 1990s that the identity of proteinaceous an important strategy employed by plants to control the movement
aquaporin water channels was confirmed, in the laboratory of of water.
Peter Agre. For this work, he received the Nobel Prize in Chemistry
in 2003. A key experiment that demonstrated the function of
a cloned aquaporin involved expressing the protein in Xenopus WATER MOVES THROUGH THE
laevis oocytes. When the aquaporin-expressing cells were trans- SOIL–PLANT–ATMOSPHERE CONTINUUM
ferred into a solution of lower ionic strength, they rapidly took up An average maize (Zea mays) plant transpires 2 to 4 liters of
water and burst within minutes. Shortly after Agre’s studies were water per day, which means that it lifts that much water up out of
completed, plant aquaporins were similarly shown to increase cell the soil, through its body, and out of the pores in its leaves,
permeability to water. without any expenditure of its own energy. A tree may lift a few
Aquaporins are major intrinsic proteins (and also membrane- hundred liters of water up to 100 m from the soil in its transpiration
intrinsic proteins) that are abundant in many tissues. In plants, there stream. These feats are astonishing, and even as late as the end of
are several families of aquaporins, including the plasma membrane the 19th century, scientists disputed the veracity of the cohesion–
intrinsic proteins (PIPs) and tonoplast intrinsic proteins (TIPS), that tension model, as it was too difficult to imagine as a viable
facilitate water movement through the plasma membrane and explanation. The more we learn about water movement in plants,
tonoplast (vacuolar membrane), respectively. Aquaporins are the more it becomes evident that the anatomy and physiology of
membrane-embedded proteins in which six membrane-spanning all stages of the water route have been adapted for efficiency and
domains are arranged to form a gated channel. They are present durability.
in membranes as tetramers, but each monomer folds into a The flow rate of water in the plant is determined by the water
separate channel. Water is not the only molecule diffusing through potential gradient and the conductance of the pathway. Factors
aquaporin channels; several of them represent important mem- that determine the water potential difference include the water
brane-selective pathways for small solutes, such as boron or silicic content of the soil and the water vapor concentration in the
acid, and CO2. There are more than 30 genes encoding aquaporins atmosphere, both of which are related to water potential. Other
in plants, which fall into four or more families. Some aquaporin gene things being equal, plants in arid environments tend to lose
families are only found in bryophytes, suggesting that they have water faster because of the very low water potential of the air
been selectively lost during tracheophyte evolution. surrounding them.
A functional role for aquaporins in plant–water relations has Conductance is determined by many different factors. At the
been demonstrated many times. Aquaporins are especially end of the pathway, the guard cells of the stomata determine
sensitive to inhibition by mercury, and so treatment of a plant whether or not the plant transpires intensely. When the stomata
with mercury gives an indication of the flux through aquaporins are fully closed, leaf surface conductance is very low (only
(although mercury is not totally specific). In various species, mercury a residual conductance exists that is attributable to leakage past
reduces root conductance by 30 to 90%. Antisense and mutational the guard cells and through the hydrophobic cuticle), and little
approaches are also useful; antisense inhibition of either the PIP1 water leaves the plant. When the stomata are wide open,
or PIP2 subclass can reduce root hydraulic conductance by transpiration is governed by the vapor pressure gradient
50%, and loss of function of pip2-2 decreases conductance by between the leaf and the surrounding air. The extent to which
30%. In Arabidopsis thaliana, antisense inhibition plants com- the water potential of the leaf is affected by transpiration is
pensated for this decrease in conductance by increasing their governed by the hydraulic conductances of the plant, including
root mass. The cells of plants expressing an antisense construct the conductance through the root, largely mediated by the
that interferes with PIP1 expression are less water permeable, activity of aquaporins, the conductance of the xylem conduits,
and the plants wilt more quickly in response to watering with a determined by their anatomy and the presence or absence of
high-osmolarity solution. Other studies suggest that the aqua- water vapor obstructions, and the conductance of the living cells of
porins have diverse roles in plant development, water flow, and the leaf, as water moves from the xylem to the stomata. All of these
environmental responses in leaves as well as roots. are under genetic control and subject to selection, and many are
Aquaporin activity is highly regulated. Many aquaporin genes environmentally responsive as well, allowing plants to optimize
show differential cell type–specific expression patterns, for example, the rate of transpirational water flow and leaf water potential in
with preferential expression in the root or the bundle sheath cells many different conditions. To better understand how plants have
surrounding the xylem. Many aquaporin genes are transcriptionally optimized water uptake and transport, we will discuss separately
regulated by light, circadian cycles, osmotic stress, or the hormone the contributions of the anatomy and physiology of the roots,
abscisic acid (ABA). Like some other plasma membrane proteins, xylem conduits, and leaves.
including auxin transporter proteins and brassinosteroid hormone
receptors, the subcellular localization of aquaporins is regulated.
PIPs have been shown to dynamically move from the internal UPTAKE AND TRANSPORT OF WATER BY ROOTS
membranes to the plasma membrane and then back to internal
membranes in response to osmotic changes. Finally, the water- For the majority of vascular plants, most of the water uptake
transporting activity of each channel is gated; that is, the pore can takes place in roots in contact with soil and is affected by many
open and close rapidly to control water movements. This occurs factors, including the soil properties and soil water content, the
through posttranscriptional modifications such as phosphorylation, architecture of the root system, the abundance of root hairs and
but there is also sensitivity to pH. Thus, aquaporin activity levels are mycorrhizal fungi, and the conductance of the root system.
 January 2014 5

Soil Properties Affect Soil Water Potential and the Transfer its dissolved ions can pass through the porous cell wall matrix (the
of Water into the Root apoplast) of the root cortical layers by bulk flow. Alternatively, it can
pass through a plasma membrane and into the cytoplasm and then
The ability of soil to retain water and to make it available to roots move from cell to cell through plasmodesmata; this is known as the
depends on many factors. One of the most important is the symplastic route, because water moves through living cells.
particle size of the soil. Soils are classified into different types, but Alternatively, the water can take a transcellular route, in which it
most soils are very heterogenous mixtures of particles of various crosses plasma membranes many times while moving through
sizes. Clays have very fine particles (;0.002 mm) predominating, cell walls and cytoplasm. The plasma membrane is a selective
loam has abundant intermediate-sized particles, and sand has barrier through which water and some small molecules can pass
particles as large as 2 mm. Field capacity is a term that defines but that acts as a barrier to most larger or charged molecules.
how much water the soil can hold before additional water runs off. Water moves across the plasma membrane by simple and
Field capacity is determined by the particle size distribution, with facilitated diffusion, which are slower processes than bulk flow.
smaller particles being more effective at holding water. Particle Thus, the plasma membrane is a source of much of the radial
size and particle charge also determine how much water is resistance of water entering the root, and the facilitation of water
tightly bound to the soil and, therefore, not available to a plant. movement by aquaporins is extremely important in the control of
The wilting point of a soil can be determined experimentally as water movement into the plant.
the soil water content below which most plants (e.g., wheat or Because it has not passed through a permeability barrier,
maize) cannot recover from wilting. Besides particle size, the water moving through the apoplastic pathway toward the stele
water-holding capacity of a soil is affected by organic matter carries in it potentially harmful dissolved ions. The Casparian strip
(e.g., humus) and the presence of mycorrhizal fungi and other of the endodermis, the cell layer immediately outside of the stele,
biological matter. is a lignified hydrophobic matrix that largely blocks the passage of
water through the apoplastic space and forces it to cross the
Root Architecture Affects the Volume and Depth from plasma membrane, thus selectively filtering it. It is named after
Which Water Can Be Extracted Robert Caspary who recognized its significance and who was
a contemporary and correspondent of Charles Darwin. Many
Roots have several functions, including water uptake, storage, older roots produce a similar barrier layer called the exodermis
anchorage, and nutrient uptake. Each of these functions is affected in the root cortex. These barriers are substantial sources of
by the distribution and abundance of the surface area of the root. resistance to water movement into the xylem-borne transpira-
Root architecture, which describes the distribution, length, angle, tion stream.
and number of individual roots and root hairs, is a highly plastic
phenotype. Genetically identical plants will produce very different
root systems when growing in different environments. In general,
FLOW OF WATER THROUGH THE XYLEM
when water is limiting to plant growth, shoot growth will be greatly
diminished, but root growth, particularly primary root elongation, The evolution of tracheary elements, the water-conducting cells
much less so. However, different kinds of drought can produce of tracheophytes, was one of the most important events in the
different distributions of root growth. When water is withheld, root evolutionary history of plants and our modern biosphere. Flow of
systems often tend to grow deeper (to reach water deeper in the water through tracheary elements is much more efficient than
soil), but when rain is regular and light, the root systems of many through living cells because (1) conduits are formed from dead
plants tend to spread out near the surface, to capture rain before it cells, so there are no plasma membranes to cross, thereby
runs into deeper soil. Roots are able to sense and grow toward reducing resistance to flow, (2) many tracheary elements tend to
water, in a process known as hydrotropism. The mechanisms are have a large diameter, which, as described by Poiseuille’s law,
still being unraveled, but perception through an unknown sensor greatly enhances flow, and (3), the thickened secondary walls of
seems to take place at the root cap, and the response involves the tracheary elements can withstand the large tensions associated
hormones ABA and cytokinin. with the cohesion–tension-based water flow. These efficient conduits
can provide much more flow of water to the photosynthetic
Root Conductance Affects Water Flow tissues than if the plant depended on transport through living
cells. Under many conditions, the rate of photosynthesis is
As described earlier, the flow of water is determined in part by the limited by the uptake of CO2 into the leaves, and CO2 uptake is
conductance (the inverse of resistance) of the conducting tissue. limited by water loss through open stomata. Therefore, a more
Conductance into and through the root system consists of two efficient water transport system means that more CO2 can be
distinct components: radial and axial conductance. Radial con- assimilated, potentially leading to a faster rate of growth. A
ductance affects water movement from the root surface to the strong selective pressure toward tracheary elements with larger
transpiration stream in the xylem, and axial conductance affects conduits can be seen from the fossil record. The earliest
movement through the xylem along the length of the root. In most vascular plants, such as Cooksonia, had tracheary element
conditions, the main limitation to water uptake is due to a limitation diameters of ;5 μm, whereas some modern vines and lianas
in radial rather than axial conductance. have tracheary elements as wide as 500 μm. As taller plants can
To reach the central vascular cylinder (stele) and the xylem within appropriate a limited light resource, the capacity for long-
it, water has to pass through the outer cortical cell layers. Water and distance vertical water transport via an efficient xylem has
6 The Plant Cell

significant adaptive value. However, as will be shown later, this affect their functions in terms of hydraulic conductance, resistance
increase in efficiency is not an unalloyed benefit. to blockage, and structural support

Tracheids: Tracheary Elements of Gymnosperms, Ferns,

Development and Differentiation of Tracheary Elements
and Lycophytes
Tracheary elements are the functional, water-conducting cells
The water-conducting cells of most gymnosperms, ferns, and
of the xylem and are either vessel elements (mostly in angio-
lycophytes are tracheids: hollow conduits largely reinforced by
sperms) or tracheids (mostly in gymnosperms, ferns, and lycophytes).
lignified secondary walls. Tracheids are best characterized from
Vessel elements and tracheids differ structurally and functionally. In
conifers, so we will restrict our discussion to these. Tracheids
addition to tracheary elements, xylem tissue can include living cells
are long cells with tapered ends. They range from less than 0.1
called xylem parenchyma and xylem ray parenchyma intimately
cm to more than 0.5 cm long and have diameters from 5 to 80
associated with the tracheary elements and, in angiosperms, fibers
μm. As water flows from tracheid to tracheid, it moves through
that are specialized for mechanical tissue support.
partially digested regions of the cell walls, known as bordered
The development and organization of primary tissues occur
pits. The pit membrane is within the bordered pit and separates
in root and shoot apical meristems, in the distal regions of the
adjacent cells. Pit membranes are not lipid bilayers or plasma
plant. Vascular tissue is specified at the meristems and during
membranes but instead are thin, water-permeable regions com-
the development of nascent leaves and lateral roots. The
posed of cellulose, which is derived from the primary walls of
development and differentiation of tracheary elements in-
adjacent cells and the middle lamella between them. In conifers,
volves three key stages: patterning, differentiation, and death.
the pit membrane has a center torus that is thick and water
Vascular tissue patterning is largely determined by patterns of
impermeable, surrounded with a margo that is highly permeable.
polar auxin flux, which induces the expression of a transcrip-
As compared with the pit membrane of angiosperms, the conifer
tional regulatory cascade leading to cell differentiation. During
pit membrane has a relatively low resistance to water flow.
differentiation, the future tracheary element cells can grow to
a very large size; tracheary elements can be 1 cm or more in
length and in some plants can reach diameters of 500 μm. Once Vessels, Vessel Elements, and Fibers: Xylem
the cells reach their final size, they lay down elaborately patterned of Angiosperms
secondary cell walls, which reinforce the primary cell walls. The
pattern of secondary wall formation is species-specific and In the xylem of most angiosperms, the functions of support and
developmentally regulated. Lignin, a polyphenolic polymer, is water transport are distributed to two cell types, the fibers and
key to the function of this secondary wall. Unlike the primary wall, vessel elements, respectively. Fibers are hollow, lignified cells
which retains malleability and water permeability, the lignified that are not normally water-conducting cells and will not be
secondary wall is rigid and much less permeable. The final stage discussed here further. The conduit of water transport is the
of tracheary element differentiation is programmed cell death vessel, which is composed of several vessel elements stacked
accompanied by lysis and degradation of the cellular contents, one upon the other. Vessel elements tend to be roughly cylindrical
leaving behind a hollow, rigid water-conducting conduit. and less tapered than tracheids. End walls are either entirely open
Many plants are able to continually produce new vascular tissue, (with no primary or secondary walls) or partially enclosed by
most commonly known as “wood,” in a process called secondary secondary walls. Thus, there is little resistance to water flow from
vascular development. The vascular cambium is the meristematic vessel element to vessel element, so the functional unit is the
tissue responsible for secondary growth. In seed plants, the multicellular vessel. Vessels can be very long, from 1 cm to more
vascular cambium is a bifascicular (meaning “two-faced”) plurip- than 1000 cm, with diameters from 15 to 500 μm. The greater
otent stem cell population that produces precursors to both xylem length of open, unimpeded vessels as compared with tracheids,
and phloem. Most gymnosperms and angiosperms, with the as well as their larger diameter, should mean that angiosperm
exception of monocots, produce vascular cambia and can tracheary elements are less resistant to flow than those in
produce secondary vascular tissues. Some plants, trees for conifers. However, the greater conductance of the conifer pits (as
example, grow extensively in girth during their lifetime. With its well as the greater total area often occupied by tracheids in
massive trunk, the giant sequoia (Sequoiadendron giganteum) conifers) compensates somewhat for the morphological differ-
is a good example of extensive secondary growth; the General ences, which leads to the question, “How does xylem structure
Sherman tree in California is considered the largest living tree affect its function?”
by volume and has a trunk diameter of 7.7 m, and the Wawona
tunnel tree was famous for having a tunnel cut into it through
which cars could drive.
Xylem Conductance

As described by Ohm’s law, the flow of water through the xylem is

Structure and Function of Tracheary Elements the product of its driving force (the difference in pressure potential
between the two ends) and the xylem conductance. As described
The structures of tracheary elements are unusual and diverse, by Poiseuille’s law, flow scales with the radius of individual
and to understand them, we must consider how their structures conduits to the fourth power, so the maximal xylem conductance
 January 2014 7

generally is higher in species with larger diameter conduits. The diameter of the conduits also contributes to resistance or
Conductance can be quantified in living tissues in several ways. sensitivity to embolism. For example, species with an average
Conductance can be measured in excised stem segments by conduit diameter of greater than 30 μm are particularly
measuring the amount of water that moves through it (by susceptible to cavitation induced by freezing and thawing. When
weighing the efflux over a period of time) for a given pressure water in the xylem freezes, dissolved air becomes less soluble
gradient or it can be measured in vivo by applying a heat pulse to and comes out of solution as gas. Upon thawing and the
a branch and determining the time it takes to reach another point reestablishment of tension in the xylem sap, the gas bubbles can
downstream to determine the flow rate, which, when divided by either dissolve again or expand and form an embolism. Smaller
the pressure gradient, provides the conductance. bubbles usually dissolve, larger ones usually embolize. Narrow
Xylem conductance is affected by the anatomy of the tracheary conduits prevent large bubbles from forming, thus conferring
elements, but nevertheless, it can vary with conditions. Recently, resistance to freeze–thaw-induced embolism. Consequently,
xylem conductance has been shown to be sensitive to the ionic large conduits are rare in high-latitude or high-elevation species,
composition of the xylem sap; specifically, the presence of cations in spite of the greater conductivity they provide. In contrast,
in the xylem sap increases pit membrane conductance relative to coniferous species, with their small-volume tracheids, are more
pure water. Imaging of the pit membranes with atomic force resistant to permanent embolism and thus well adapted to cold
microscopy revealed that their thickness decreases when cations regions. However, due to the important roles of pit membranes in
are present, which increases their conductance. Other studies resisting cavitation and embolism, the relation between conduit
suggest that this ionic effect is adaptive and functionally relevant, size and cavitation resistance is not simple.
although to what extent it is under the control of the plant is not Many methods have been developed with which to measure
certain. embolism formation. These include the quantification of acoustic
emissions (sounds) that occur with cavitation and imaging methods
involving scanning electron microscopy or x-ray–based computer-
“The Vulnerable Pipeline”: Cavitation and Embolism assisted tomography (these are described in more detail under
Formation Methods Used in Plant–Water Relations Research). One of the
more widely used methods is to determine the conductivity of
The cohesion–tension mechanism of water transport requires a tissue at several different water potentials and from these data
little or no internal energetic expenditure. There is an associated construct a vulnerability curve or a percentage loss of conductivity
cost, however, which is that the system is vulnerable to potentially curve. Percentage loss of conductivity curves provide useful
catastrophic embolism formation. This occurs when air enters information about the sensitivity of the plant to environmentally
through the pit membranes from an adjoining air-filled conduit, induced embolisms, which generally correlate with where the
a phenomenon referred to as cavitation. Because of the tension plant can survive. For example, plants endemic to humid areas
in the surrounding sap, the air expands to form an air-vapor lose their hydraulic conductivity at higher (less negative) water
blockage that breaks the water column, a condition known as potentials than those from dry regions.
embolism. A fully gas-filled conduit is no longer able to con- The conductance of a plant can decrease over a growing
tribute to water flow. Under some conditions, including drought season as more and more xylem vessels are blocked by em-
and freezing temperatures, water flow can be extensively bolism. In some species, root pressure can refill the embolized
impeded by embolized xylem, even to the extent of the death vessels (as described further below). Gymnosperms and angio-
of the plant. sperms are able to replace nonfunctional xylem by the action of
The anatomy of the pit is important in resistance to drought- the vascular cambium, which produces new xylem. Most ferns
induced cavitation and in the resistance to embolism spread. and lycophytes are not able to generate secondary xylem.
Drought causes increased tension in the xylem conduits, because Interestingly, studies have demonstrated that ferns have a very
water is held at greater tension in the soil and transpirational conservative strategy to prevent embolization of their vascular
demand will increase this tension as water flows across much tissues. Relative to the maximum xylem tension that each can
steeper water potential gradients than would develop in a well- endure, fern stomata close in response to less xylem tension
watered plant. Increased tension most likely induces cavitation to than angiosperm stomata; they stop transpiration, and therefore
occur by air seeding from an adjacent gas-filled tracheary photosynthesis, before there is a great risk of xylem failure due
element or the atmosphere at the pits between conduits. Pits to embolism formation. In other words, if you can’t replace it,
that resist air seeding can reduce drought-induced losses of you’d better take care of it!
hydraulic conductance. Some xerophytes produce vestured pits, Thus, in xylem construction, there has to be a balance between
which have an elaboration of the secondary wall filling the pit hydraulic efficiency and safety from embolism. These safety–
cavity and which are thought to aid in resisting cavitation. The efficiency tradeoffs are found at the level of the pit, the conduit, and
thickness and size of the pit membrane, as well as its interactions the whole plant. Essentially, plants cannot do everything well.
with adjacent secondary cell walls, also can affect resistance to Hydraulically efficient species such as water birches are susceptible
embolism spread. The torus-margo arrangement of bordered pits to embolism, while drought-resistant species such as Ceanothus
found in conifers can be particularly efficient at resisting the shrubs have a very low hydraulic efficiency but high cavitation
spread of embolisms between cells. Anatomies that resist embolism resistance.
formation or spreading are found in plants that regularly experience A lively debate is taking place right now about the extent to
drought or freezing stress. which embolisms can be refilled when the xylem is under
8 The Plant Cell

tension. We refer the reader to the articles cited in Recom- Xylem under Pressure: The Roles of Root and Stem
mended Reading for an elaboration of the arguments for and Pressure on Water Movement
against this model.
Early plant physiologists wondered if the flow of water in the
xylem was a result of pressure in the roots forcing water upward.
Although Hales and others demonstrated that this is not the
A FEW EXAMPLES TO ILLUSTRATE WATER MOVEMENT
case most of the time, there are cases in which root or stem
IN XYLEM
pressure is associated with water flow.
Mangroves and Ultrafiltration of Seawater

Like Stephen Hales, Per Scholander studied the physiology of Refilling of Grapevines by Root Pressure
both plants and animals, and much of his work was on the diving Grapevines (Vitis vinifera) have long been used as a model system
physiology of marine mammals. We remember him for his for the study of plant–water relations. Not only are they economically
studies of xylem tension, using the Scholander pressure bomb important, but they also have some of the largest diameter vessels
method, in plants from several habitats, including mangroves. of any plants. Large vessels mean that water flows efficiently, but
Mangroves are diverse plants from many different families that they also mean an increased vulnerability to freezing-induced
share the feature of growing with their roots in brackish water or cavitation. Because of this, grapevine vessels are empty (air filled)
seawater. Mangroves must obtain water from a salty substrate in the winter. Prior to leaf emergence, the vine vessels fill with
with a low water (osmotic) potential. This is analogous to growing water, which is accompanied by a high positive xylem pressure.
in a perpetually very dry soil. Scholander reasoned that the highly The pressure is such that a cut vine exudes copious amounts of
negative osmotic potential of seawater would pose a significant water and is said to “bleed.” Pressurized vessel filling is a short-
challenge to the ability of these plants to take in water. In 1962, he lived effect, and after the buds open and leaves form, transpiration
asked, “What balance, if any, exists between osmotic potential of begins and the vessels show a negative pressure. The spring-
seawater, roots, and leaves on one hand and hydrostatic pressure time positive pressure originates from osmotic pressure in
and osmotic potential of the xylem sap on the other?” Mangroves the root xylem, produced by the active transport of solutes.
have adapted to their environment through the development of Note that guttation, or the forcible extrusion of water from the
very low osmotic potentials in living cells (by the accumulation hydathodes at the edges of leaves, is also a product of root
compatible solutes such as proline) that allows them to develop pressure, which can occur when plants are not transpiring and
low leaf and root water potentials sufficient to pull water into the the water potential gradient nevertheless draws water into the
roots. They also must possess sufficiently strong resistance to roots from the soil.
high xylem tension. In 1964, Scholander measured xylem tension
in mangroves at about 23 to 25 MPa, much lower than that of
a “typical” plant. As Scholander observed, mangroves face “an A Sweet Enigma: Stem Pressure and Flow of Maple Sap
exaggerated version of the old problem of how sap rises in
The flow of maple sap is an interesting example of an unusual form
tall trees.” In other words, both mangroves and tall trees require
of sap movement, driven by stem pressure. Sugar maples (Acer
very low water potentials in the xylem to bring water to their
saccharum) are common in the northeast part of North America and
leaves.
have been used for centuries as a source of sugar and syrup.
During early spring, sweet sap flows from a spike, called a spile,
Does Xylem Conductance Limit the Height of Trees? driven into the trunk of a tree. The mechanism of sap flow has long
puzzled plant physiologists and relies in large part on the unusual
Until now, we have ignored the effect of gravity on water potential. anatomy of the trees. In sugar maple wood (and in many other
However, for very tall trees, we must include it. The effect of trees), the structural fibers that are interspersed with the vessels are
gravity on water potential is to lower it by ;0.1 MPa per 10 m of air filled. Sap flows only under very specific conditions, which gives
height. As water moves up the tree, the weight of the water clues to its mechanism. Sap flows from barren, leafless trees,
column below it increases, and the water potential decreases with indicating that it is not a transpiration-driven process. In contrast
increasing distance up the tree. Therefore, for a tree of 100 m, an with grapevines, if a maple branch is cut off, the sap flows out of the
additional 1 MPa of tension is experienced in the xylem at the tree distal “leaf” end rather than the proximal “root” end of the cut; this
crown as compared with the roots. Koch et al. (2004) showed that suggests that it is not root pressure that drives sap flow. In fact, an
the growth rate and photosynthetic rates are much lower for isolated segment of branch without any roots can release sap. Also
leaves at the top of a coast redwood tree (Sequoia sempervirens) unlike grapevines, maple sap flow is temperature dependent and
than lower down the tree and proposed that trees taller than ;120 occurs when below-freezing nights are followed by above-freezing
m are growth-limited by hydraulic limitation. However, others days. An isolated branch takes up water into its cut end when the
have argued that other factors are equally if not more important temperature is below freezing and exudes it when the temperature
for growth limitation, such as energetic tradeoffs for continued is above freezing. This cycle argues against sap flow being driven
structural investment, nutrient limitation, and repeated dieback, by the change in volume of freezing water, because water expands
as well as various combinations of these. These discussions are when frozen. Instead, the current model suggests that the freezing
indicative of the inherent difficulty of measuring and observing temperatures draw water into the air-filled cavity of the fibers, which
water flow in the xylem of woody plants. then freezes, compressing and pressurizing the air. When the ice
 January 2014 9

thaws, the pressurized air forces the water back into the vessels more heavily. In some nondeciduous conifer leaves (which are not
and down the tree trunk. The continuing sap flow probably requires as short-lived as deciduous leaves), high xylem tension can cause
additional osmotic pressure in the vessels. Again, the anatomy of the leaf tracheids to collapse reversibly, which by sealing off the
the maple supports this possibility, because the vessels are unusually stem xylem system from the transpirational pull may protect it
devoid of pits connecting them to the fibers, which raises the from embolism.
possibility that sucrose can move into the vessels but not out of
them, leading to an increased pressure driven by the influx of water.
Unfortunately, sap flow’s dependence on freezing temperatures Outside-of-Xylem Conductance
means that climate change is expected to restrict sap production in
In angiosperms, the leaf vasculature is surrounded by specialized
traditional sugar maple–growing regions.
mesophyll cells called bundle sheath cells. In C4 photosynthetic
plants such as maize, these cells have been thoroughly studied for
their contribution to CO2 fixation, but they are found in C3 plants
MOVEMENT OF WATER FROM XYLEM TO LEAF CELLS TO as well. Water leaving the xylem passes through these cells,
THE STOMATAL PORE which are thought to vary in conductance with environmental
conditions. Furthermore, there is some evidence suggesting that
The movement of water within the leaf takes place as liquid
the bundle sheath cells may have a similar role to that of the
water moving through the xylem and the living leaf cells and as
endodermis in controlling the flow of solutes into the leaf tissues.
water vapor within the intercellular spaces of the leaf. The rate of
The hydraulic conductance of the radial pathway from vein to
flow is determined in part by the conductance of each of these
stomatal pore may be at least partially regulated by ABA and by
segments of the pathway: within the vascular tissues, through
drought signals, which are likely to be mediated at least partially
the nonvascular tissues (via apoplastic, symplastic, and trans-
through aquaporin activities.
cellular pathways), and stomatal conductance.
The flow of water from the xylem to the mesophyll requires that
the water potential of the mesophyll cells is lower than that of the
xylem. Xylem water potential is primarily determined by tension,
Leaf Vein Anatomy and Conductance whereas mesophyll cells have a positive turgor pressure. If the
mesophyll cells had the same osmotic potential as the sap in the
Within the leaf, the xylem provides an efficient and low-resistance
xylem, water would flow backward into the xylem. The fact that it
pathway for water movement. In most leaves, the vein pattern is
does not is due to the lower osmotic potential of the mesophyll
such that the photosynthetic cells are all located very close to a
cells. For a well-watered plant, the osmotic potential of mesophyll
vein, ensuring them both a steady supply of water and a relatively
cells can be on the order of 20.5 to 21.5 MPa. However, for
short pathway to return photosynthate to the vascular tissues.
plants adapted to low-water environments or plants under drought
Vein density is notably greater in angiosperms than in non-
stress, the water potential of the xylem can be considerably lower,
angiosperms. The fossil record shows that increased vein density
23 MPa or lower, so the mesophyll cells must compensate by
appeared ;100 million years ago, and this higher density may be
accumulating additional solutes, such as ions or organic solutes.
essential to support the high rates of transpiration and photosyn-
Thus, the leaf adjusts its osmotic milieu to ensure that the water
thesis of which angiosperms are capable. Veins in dicots tend to
potential of the xylem sap is not lower than that of the living leaf cells.
form branching reticulated networks, whereas in monocots, there
is a tendency for parallel veins or parallel secondary veins branching Stomatal Conductance
off a single midvein; these redundancies ensure that there are
alternative pathways for water to flow if one gets damaged or Stomata are the tiny openings in the leaf epidermis and the adjacent
embolized. pairs of guard cells that act as gatekeepers of transpiration/gas
Because of the relatively lower conductance of mesophyll cells exchange. (In some species, the stomatal complex includes
as compared with xylem, conductance is negatively correlated with additional neighboring cells that support the activities of the
distance from vein to evaporating surface, so the greater vein guard cells.) When conditions are right for the guard cells to
density of angiosperms means shorter vein-to-stomata distances open, ions are pumped into the cytoplasm, which leads to the
and is correlated with higher maximum photosynthetic rates. Some movement of water into the cell by osmosis. Because the guard
gymnosperms have additional non-xylem-conducting tissues called cells have a rigid inner wall and radial bands of cellulose, as they
water-conducting sclereids that facilitate water movement through increase in volume their outer radial wall expands, which leads
the mesophyll. Aquaporin activity in the leaf mesophyll can also to the pair of cells moving away from each other, opening the pore
increase the conductance of the pathway and has been shown to that they cover. When the stomatal pore is open, CO2 enters and
be regulated by many environmental parameters, including light, water vapor is released by diffusion. When water needs to be
temperature, and drought stress. conserved, plasma membrane–localized ion channels open,
Several studies have suggested that one function of the xylem ions and water flow out, and the guard cells lose turgor and
within leaves may be to protect the xylem in the stem. For relax together, covering the pore and blocking transpiration.
example, in some deciduous plants, the xylem of the leaves is less Most guard cells are extremely sensitive to the hormone ABA,
resistant to cavitation than that of stems or branches, suggesting which promotes stomatal closure and inhibits stomatal opening.
that the conduits in these leaves may act as disposable “fuses” Downstream of ABA are a family of receptor proteins that operate
that help protect tissues in branches that the plant has invested in as dimers and, when bound to ABA, regulate both transcription
10 The Plant Cell

factor and ion channel activities. The immediate effect of ABA mortalities of forest trees could further exacerbate the rate of
on guard cells is to open potassium and anion channels, carbon emission and climate destabilization, potentially leading
releasing ions from the cytoplasm, followed by water, and pore to a positive feedback cycle that we must avoid.
closure. Decreases in stomatal aperture and transpiration can be
recorded within minutes of ABA application to a leaf or epidermal
strip. SUMMARY AND ONGOING RESEARCH
ABA can be synthesized in guard cells and their neighbors in
response to decreasing relative humidity. ABA can also be carried The cohesion–tension model states that water moves through
through the transpiration stream from drying roots to guard cells. plants without any mechanical pumping required, and this can be
Furthermore, ABA can affect transpiration by altering the conduc- demonstrated by a simple clay pot on a tube. When you consider
tance of nonstomatal cells of the leaf and root. Other studies have how much energy would have to be extended to move water up
shown that a hydraulic signal can contribute to guard cell responses a 100-m tree if pumping were involved, this feat seems remarkable,
directly. The relative importance of hydraulic signals and ABA is still and acceptance of the cohesion–tension model was slow.
being debated; some studies have suggested that guard cells of Nevertheless, it is now the widely accepted explanation for water
earlier emerging plants such as ferns and lycophytes are insensitive uptake and transport in plants. Plants are much more complicated
or less sensitive to ABA than are those of angiosperms. Interestingly, than clay pots on sticks, and the results of millions of years of
the genomes of these earlier emerging plants reveal that genes evolutionary selection underpin every step of the water-flow
required for ABA responses are present. How and if hydraulic signals process. From the far ends of the roots to the highly sensitive
integrate with ABA signals in the regulation of water transport guard cells, living plant cells sense and respond to water, allowing
continue to be fascinating questions. Stomata also respond in an them to maximize CO2 uptake while minimizing dehydration injury
adaptive fashion to other environmental signals. For example, and damage to their vulnerable conducting tissues. Millennia of
guard cells respond to light (opening) and dark (closing) and low evolution have produced highly efficient tracheary elements, with
humidity (closing) stimuli, all of which promote a greater ratio of anatomical features that can be genetically and perhaps physio-
CO2 molecule capture in photosynthesis per molecule of water logically fine-tuned to provide greater hydraulic conductance or
loss. embolism resistance. The living tissues of the shoot and root are
more resistant to water movement than the hollow conduits of the
xylem but also provide more opportunities for regulatory controls.
DROUGHT, HYDRAULIC FAILURE, AND WHAT There is still much that we do not understand yet about water
IT ALL MEANS uptake and transport, and the field is as rich with conflicting models
and hypotheses as it was when the cohesion–tension model was
Plants tend to have a system for water uptake and transport that put forth more than 100 years ago. Exciting new methods and
is well suited for the niche they occupy and for competing with technologies such as high-resolution computed technology can test
species occupying similar niches. Fast-growing species in moist established models in new ways and pave the way for new models.
environments may have large conduits and numerous stomata The genetic resources of Arabidopsis as well as other model
to support a high rate of transpiration and carbon uptake but that species, and the power of transcriptomics, proteomics, and
makes them vulnerable to drought stress. Arid-climate species metabolomics methods, are being applied toward these ques-
may have a more conservative approach to transpiration and gas tions. Our forests may be the most important beneficiaries of
exchange that limits their competitiveness in moist environments these studies, as long-lived, tall trees are particularly vulnerable to
but allows them to tolerate arid ones. Root and shoot size and the kinds of dramatic changes in rainfall patterns and water
architecture can also be optimized to support the balanced needs availability that are being postulated in coming years. As the
for water uptake and photosynthesis in a given niche. Most authors of an influential paper (Choat et al., 2012) point out, “If the
species have some range of tolerances to account for seasonal tight link between embolism resistance and water availability is
and annual variations in environmental conditions. However, the product of natural selection over many generations and
prolonged conditions of reduced water availability (drought) are adaptation is limited by a long generation cycle of perennial
often lethal. In some cases, particularly managed and agricultural plants, then the rapid pace of climate change may outstrip the
systems, drought can be ameliorated by irrigation. In others, capacity of populations to adapt.”
particularly forest and natural systems, a lack of rainfall can push
a plant beyond its tolerances. Drought lowers the water potential
of the soil, leading to lower water potentials in the xylem. Beyond METHODS USED IN PLANT–WATER
a threshold level of hydraulic tolerance, depending on the interplay RELATIONS RESEARCH
between xylem anatomy, stomatal responsiveness, and root
uptake efficiency, a droughted plant can experience hydraulic Some of the liveliest discussions that take place among plant
failure, which may result in death. In recent years, there have been scientists are those about the relative merits of various methods for
documented increases in drought-induced vegetation mortality measuring a plant’s hydraulic properties. Simply put, there is no
associated with climate change. This condition is often exacer- perfect method, as these properties are extremely challenging to
bated by the increased vulnerability of drought-stressed trees to measure. Without getting too deeply into the controversies, we will
further damage by pests such as bark beetle and opportunistic introduce a few of the methods commonly used and refer the
pathogens. Because forests store vast amounts of carbon, widespread reader to Recommended Reading for more thorough discussions.
 January 2014 11

Transpiration, Water Uptake and Release, and Water longer the time the pulse takes to reach the sensor, the slower
Potential the rate of flow in the xylem. A more sensitive variation of this
technique involves adjusting the rate with which heat has to be
A simple method to measure the movement of water at the whole- introduced to the stem in order for a constant temperature to be
plant level was developed by Stephen Hales in the 1700s. This maintained. As the flow rate increases, the rate of heat addition has
involves weighing a plant in a sealed pot at various times, with the to be increased as well. Dyes or tracers can be used instead of heat.
loss of mass being attributed to the movement of water from the
pot to the atmosphere through the plant. This method is still widely
used today, particularly for small, pot-grown plants like Arabi- Conductance
dopsis and rice (Oryza sativa), but it has several limitations in Hydraulic conductance can be measured by measuring the flow
addition to the need for the plant to be grown in a pot. In through a branch, branch or root segment, or root system, driven by
particular, the method measures whole-plant flow but does not various pressure gradients. Tyree and colleagues developed a high-
differentiate which segments of the plant are responsible for any pressure flow meter method for measuring root conductance in situ,
variation in flow rate. in which a root or root system is excised from the shoot (but left in the
Alternatively, a porometer can be used to measure transpiration soil) and a pressurized water source is attached to the cut end. The
or stomatal conductance in real time. A porometer has a chamber flow rate at which water enters the root system can be plotted
that can be clamped onto a leaf, and the amount of water vapor against the pressure applied to determine conductance. In Tyree’s
that moves from the leaf into the device is measured by a change in method, the water was forced from the root base (near the shoot) to
chamber humidity. Porometers are portable and widely used in the the distal end (i.e., rootward), the opposite direction to the normal
field. A similar principle is used in thermocouple psychrometers, water flow. Root conductance also can be measured by placing
which determine the water potential of a sample placed into a root system into a pressure chamber and measuring water flow
a chamber. Psychrometers can be used in determining the water from the cut surface (i.e., shootward flow).
potential of plant or soil samples. Hydraulic conductance or conductivity across a cell membrane
Transpiration can also be indirectly measured by a change in can be measured by stripping off the cell wall, immersing the
leaf temperature, as the water evaporation withdraws heat protoplast in a solution of defined osmotic potential, and measuring
energy from the leaf. Temperature changes can be measured the change in cell volume. Because water movement across a
through infrared imaging of single plants or leaves or of entire membrane occurs largely through aquaporins, this method is useful
fields or ecosystems using satellites. in determining aquaporin activity.
A potometer can give an indirect measurement of transpira-
tion rate by recording the amount of water drawn into a cut
branch that is immersed in water; the name means “drink meter,” Cavitation and Embolism
and it records how much the plant “drinks.” The simplicity of this Vulnerability curves can indicate how well a plant tissue resists
method makes it a popular one for school science laboratories. cavitation at various xylem tensions. Vulnerability curves are
determined by measuring the flow through an excised stem
segment at progressively lower xylem tensions. As the tension
Pressure and Tension increases, flow will progressively decrease due to embolism
Dixon and Joly (1895) and later Scholander (1960s) pioneered the blockages. To determine the maximal, 100% flow, the embol-
use of “pressure bombs” to measure xylem tension. Cut branches ized vessels can be forcefully refilled with high-pressure water.
or leaves are placed into sealed chambers so that only the cut Various methods can be used to lower the xylem water potential,
surface protrudes. Because the xylem system is under tension, including progressively drying the segment to lower the xylem
water retreats from the cut surface into the severed branch or leaf. water potential or spinning the segment in a centrifuge, and
The balancing pressure applied to the chambered tissues that is conductivity is measured at several water potential values. These
sufficient to force the xylem stream back to the level of the cut “percentage loss of conductivity” curves are widely used to
surface can be interpreted as the negative of the xylem tension if compare the hydraulic properties of various plants and plant
the leaf was not transpiring. Another method to estimate tension is parts. The water pressure at which a segment embolizes can also
to measure the diameter of the stem, with more tension leading to be measured by plotting the number of cavitations (monitored by
a narrower stem. More recently, a pressure probe method has sound) occurring at various water potentials.
been developed that measures pressure or tension within a tissue Imaging methods have been developed that reveal the
by impaling individual cells with a fluid-filled needle and measuring presence of embolisms in living tissues. A relatively simple
the force exerted on the needle. approach is to cut a segment and put the cut end into a dye,
which labels the actively transpiring conduits. Another method,
cryoscanning electron microscopy, involves freezing stem seg-
Flow ments and slicing them while frozen. Air-filled (embolized) conduits
are hollow in the resulting sections, whereas water-filled ones
The flow rate of sap within a plant can be measured by heat-flow remain filled with ice, although there is some concern about the
measurements. Heat is introduced into a stem or branch, and extent to which sample processing introduces artifacts.
the amount of time required for the heat pulse to be detected at More recently, medical imaging techniques have been adapted
a sensor farther along the transpiration stream is measured. The for use in plants. Magnetic resonance imaging can distinguish
12 The Plant Cell

water from air, but the resolution of this method is insufficient to Dixon, H.H., and Joly, J. (1895). On the ascent of sap. Philos. Trans. R.
resolve single conduits. High-resolution computed tomography is Soc. B. 186: 563–576. doi:10.1098/rstb.1895.0012.
a noninvasive imaging method that can resolve fine structures Hales, S. (1727). Vegetable Staticks: Or, an Account of Some Statical
Experiments on the Sap in Vegetables. (London: W. and J. Innys).
and that has been used in the past few years to image embolism
Kramer, P.J. (1974). Fifty years of progress in water relations research.
spread and repair in grapevines. See Cochard et al. (2013) for
Plant Physiol. 54: 463–471. doi: 10.1104/pp.54.4.463.
a discussion of the reliability of these different methods.
Mansfield, T.A., Davies, W.J., and Leigh, R.A. (1993). The transpiration
Understanding plant responses to water limitation is complex, stream. Introduction. Philos. Trans. R. Soc. Lond. B 341: 3–4.
and uptake and transport is just part of the picture. In Part 2 of this doi:10.1098/rstb.1993.0085.
article, we examine how water limitation affects photosynthesis Richter, H., and Cruiziat, P. (2002). A brief history of the study of water
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