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Journal of Experimental Botany, Vol. 53, No. 378, pp. 2225±2237, November 2002
DOI: 10.1093/jxb/erf071
Chilling root temperature causes rapid ultrastructural
changes in cortical cells of cucumber (Cucumis sativus L.)
root tips
Seong Hee Lee1, Adya P. Singh1,3, Gap Chae Chung1, Yoon Soo Kim2 and In Bae Kong1
1
Agricultural Plant Stress Research Centre (APSRC), Division of Applied Plant Science, College of Agriculture,
Chonnam National University, Gwangju 500-757, South Korea
2
Department of Forest Products and Technology, College of Agriculture, Chonnam National University, Gwangju
500-757, South Korea
Received 7 January 2002; Accepted 1 July 2002
Abstract
Examination of root tips from cucumber (Cucumis
sativus L.) seedlings grown at 8 °C for varying periods ranging from 15 min to 96 h, showed marked
changes in the ultrastructure of cortical cells within
only 15 min of exposure. Greater parts of the cortex
were affected with longer periods of exposure, but
the sequence of morphological changes in cell
components was similar to that found for the roots
exposed for 15 min. The effect of chilling injury
included alterations in cell walls, nuclei, ER, mitochondria, plastids, and ribosomes. The extent of
alterations varied greatly among cells, moderate to
severe alterations to cell components being observable among adjoining cells. The measurements of
root pressure using the root pressure probe showed
a sudden, steep drop in the root pressure in
response to lowering of the temperature of the bathing solution from 25 °C to 8 °C. These observations
are discussed in the light of the information available
on the ultrastructural and biochemical characteristics
of the effect of cold exposure in chilling-sensitive
plants.
Keywords: Chilling injury, cortical cells, Cucumis sativus
roots, transmission electron microscopy, root pressure.
Introduction
Cucumber (Cucumis sativus L.) is an important crop plant
for many parts of the world, Asian countries in particular.
3
However, this plant is sensitive to temperatures below
about 15 °C, and massive economic losses can occur if the
temperature suddenly falls below this level during the
growing season.
The effect of cold temperatures on the ultrastructure
of plant cells has been studied for a long time, and
recently reviewed (Kratsch and Wise, 2000). The
extent of alterations in cell components appears to be
related to the severity of the cold temperatures and the
length of exposure. Generally, agricultural plants grown
in tropical and sub-tropical regions are susceptible to
chilling damage at temperatures ranging from 0±15 °C.
Plant response to cold temperatures appears to be both
species- and tissue-speci®c. Ultrastructural studies suggest that cold-temperature-related changes involve a
wide range of cell components, and plastids are one of
the most extensively investigated organelles. Plastids
and thylakoid membranes swell and become disorganized, thylakoids and peripheral reticulum vesiculate,
plastoglobuli or lipid droplets accumulate, and eventually the entire plastid becomes disorganized leading to
the disintegration of the envelope (Klein, 1960; Millerd
et al., 1969; Taylor and Craig, 1971; Kimball and
Salisbury, 1973; Forde et al., 1975; Moline, 1976; Niki
et al., 1978; Nesler and Wernsman, 1980; Murphy and
Wilson, 1981; Wise et al., 1983; Ishikawa, 1996).
Signi®cant alterations have also been observed in other
cell components. Mitochondria undergo distinct swelling (Ilker et al., 1976; Moline, 1976; Niki et al., 1978;
Murphy and Wilson, 1981; Ishikawa, 1996), the
population of ribosomes decreases, endoplasmic reticulum (ER) dilates, cytoplasmic membranes vesiculate,
To whom correspondence should be addressed. Fax: +82 62 5300190. E-mail: [email protected]
Published by Oxford University Press
2226 Lee et al.
the nuclear chromatin condenses forming clumps,
plasmalemma invaginates and there is an increase in
vacuolation and the number of membraneous vesicles
(Ilker et al., 1976; Moline, 1976; Platt-Aloia and
Thomson, 1976; Niki et al., 1978; Murphy and Wilson,
1981; Ishikawa, 1996).
Cucumber roots respond rapidly to 8 °C chilling
temperature, with a drastic reduction in water uptake
within about 10 min of exposure to this temperature
(SH Lee et al., unpublished observations), and thus
provide a valuable experimental system to study
various aspects of cell biology in relation to this and
other factors. Using the root pressure probe for
measurements of root pressure developed in the root
system (Steudle, 2000), evidence is provided for a
sudden, steep drop in root pressure in this species
within minutes of exposure to chilling temperature. The
ultrastructural observations reported here on the effect
of 8 °C temperature on cucumber root tips are part of
a detailed study to investigate tissue organization and
anatomical composition of various regions of cucumber
roots, including the root-hair-producing region where
much of the water uptake is likely to occur. The aim
is to understand the basis for the observed sudden drop
in the root pressure in cucumber in response to chilling
temperature using a combined physiological, biochemical and anatomical approach in this study.
Materials and methods
Cucumber (Cucumis sativus L.) seeds were germinated on moist
paper towels in plastic trays, in the dark, in an incubator. The
seedlings were grown for 4 d in the dark and were then exposed to
8 °C for periods ranging from 15 min to 96 h. About 1±2 mm long
segments were collected from the root tip region at 0 h (control) and
after the following periods of exposure to 8 °C: 15 min, 30 min, 1 h,
4 h, 8 h, 16 h, 24 h, 48 h, 72 h, and 96 h. The samples were processed
for light (LM) and transmission electron microscopy (TEM) as
described below.
Root tip segments were transferred to vials containing 3%
glutaraldehyde (prepared in 0.05 M sodium cacodylate buffer)
immediately after cutting with a sharp razor blade under a stereo
microscope, and ®xed overnight at 4 °C. After washing in the buffer,
the samples were post-®xed in 2% osmium tetroxide (also prepared
in 0.05 M sodium cacodylate buffer) and washed again in the buffer.
They were dehydrated in a graded series of acetone, and in®ltrated
and embedded in Spurr's low viscosity resin (Spurr, 1969).
Semi-thick (2±3 mm) and ultra-thin sections (60±80 nm) were cut
with glass and diamond knives, respectively, on an RMC MT X
ultramicrotome. Semi-thick sections were stained with 0.1% aqueous toluidine blue and examined with a Zeiss Axiolab photomicroscope. Ultra-thin sections were sequentially stained with uranyl
acetate and lead citrate and then examined with a JEOL 1010 TEM.
The root pressure of an excised cucumber root system was
measured by root pressure probe as previously described (Steudle,
2000). Once the cut end of a root segment was connected tightly to
the probe, the root pressure developed due to active uptake of
nutrients into root, and the root pressure was measured continuously
using the probe.
Results
The ultrastructural observations presented are based on the
conventional chemical ®xation of samples, which is known
to alter the morphology of some cell components. It is also
likely that cells undergoing cold-induced changes may
react somewhat differently to ®xatives compared to
uninjured cells. Therefore, the observations presented
have to be viewed with these considerations in mind.
Observations were con®ned to examining cortical cells
from a region about 1 mm behind the root apex. Cells in
this region were undergoing expansion growth as indicated
by the presence of developing as well as fully developed
intercellular spaces. Examination of semi-thick monitor
sections with LM provided early indications that changes
in root cortical cells due to cold treatment may be rather
rapid.
In roots receiving 15 min of cold exposure some areas of
the cortex stained only lightly with toluidine blue. Cells in
such regions appeared abnormal; they had a highly reduced
cytoplasmic density, and the presence of cell walls could
not be con®rmed (Fig. 1B). However, the staining of a
large proportion of the cortex was similar to that of the
cortex of control roots (Fig. 1A). Nuclei and cell walls
were easily recognized in these cells and they had a normal
appearance, judging by comparisons with cortical cells of
control roots.
With longer periods of cold exposure, greater areas of
the cortex were affected (Fig. 1C). However, damaged
areas did not appear to be proportional to the time of
exposure. Even after 96 h of exposure only about 25% of
the cortex appeared to be severely affected. However, this
estimate is based on single sections only and not serial
sections.
TEM observations of damaged and adjacent cortical
areas con®rmed that cells in the lighter areas were severely
affected, in addition to providing detailed information on
the nature and extent of alterations in various subcellular
components.
Control roots
Cortical cells from control roots (roots not exposed to cold)
displayed features typically found in a metabolically active
cell. Nuclei were large and spherical, containing fairly
disperse chromatin and a prominent nucleolus (Fig. 2A).
Nuclei were more or less centrally located in the cells. A
relatively dense cytoplasm contained numerous mitochondria and variously shaped plastids, which had a stroma
denser than that of mitochondria and were thus readily
distinguishable from them even at low magni®cations
(Figs 2A, B, 3A). The mitochondria had a moderately
dense stroma, which contained ribosomes, DNA and welldeveloped cristae (Fig. 6A, B). The two membranes of the
mitochondrial envelope were parallel and evenly spaced
(Fig. 6A, B). The plastids contained few internal mem-
Chilling injury in cucumber root 2227
branes which were not readily distinguishable because of
the high density of the stroma (Fig. 6A). Also, the plastid
envelope, which consisted of two parallel membranes, was
not easily resolvable because of the high density of the
plastid stroma (Fig. 6A, B) and the pleomorphic nature of
the plastid form (Figs 2A, 3A). The cytoplasm also
contained abundant cisternae of rough ER, which were
dif®cult to discern at low magni®cations, mainly because
of the high density of the cytoplasm (Fig. 2A, B). As seen
at high magni®cations, ER cisternae were long, with a
uniform intracisternal spacing between the limiting membranes, and were studded with ribosomes (Fig. 6A).
Polysomes were common also in other parts of the
cytoplasm.
The plasmalemma appeared to be closely associated
with the cell walls, which were thin and somewhat
sinuous in the intercorner regions of the cells (Fig. 2A,
B). The cell walls were thicker in the cell corner
regions, particularly in those cells which had developed
intercellular cavities or were in the process of doing so
(Figs 2B, 3A). Initially, vacuoles were small (Fig. 2A),
but they progressively enlarged mainly through the
coalescence of adjoining vacuoles, and this process
appeared to coincide with the development of intercellular cavities (Figs 2B, 3A). The intercellular
cavities were formed mainly by the dissolution of the
intercellular material in the cell corner regions, as
shown by the presence of reticular, vacuolate and
dense materials in these areas (Fig. 2B).
Cold-treated roots
Chilling injury to cortical cells of exposed roots was
apparent as early as 15 min after exposure (Fig. 1B). Injury
in these samples varied greatly in severity, ranging from
changes in a few cell components to almost complete
disintegration of the cytoplasm. Longer exposure periods
caused greater parts of the cortex to be affected, as seen in
the light micrograph illustrated (Fig. 1C). Therefore, the
following description is primarily based on the features
considered to represent sequential injury, and not necessarily on the sequence of the exposure time. Also, roots
exposed to all periods had cells showing signs of injury as
well as those which appeared to have normal ultrastructure. This feature is illustrated in Fig. 3B, where cells with
early injuries can be seen next to those without any
apparent injuries.
Fig. 1. (A) Transverse section through a control root tip. Cortical
cells show features typical of metabolically active cells. They have a
dense cytoplasm and large nuclei containing a prominent nucleolus.
LM. Bar=100 mm. (B) Transverse section through a root tip exposed
to cold temperature for 15 min. A small part of the cortex has stained
lightly, and shows signs of cell damage (arrow). LM. Bar=100 mm.
(C) Transverse section through a root tip exposed to cold temperature
for 48 h. A relatively large part of the cortex has stained lightly and
shows signs of cell damage (arrows). LM. Bar=100 mm.
Early stages of chilling injury
A noticeable increase in the density of cytoplasm, and
distention of endoplasmic reticulum (ER) were early
symptoms of injuries to cells (Figs 3B, 4A). At low
magni®cations ER cisternae were recognized with dif®culty in the cortical cells of control roots (Figs 2A, B, 3A),
whereas they were easily recognized even at the early
stages of cell injury because of the presence of wider,
2228 Lee et al.
variable intra-cisternal spaces in them (Figs 3B, 4A). The
stroma of both mitochondria and plastids became more
translucent and granular, as viewed in low magni®cation
images (Fig. 4A). Both nuclei and nucleoli became
irregularly shaped (Fig. 4A). Cell walls were thinner
than in the control material and were distorted (Fig. 4A).
Fig. 2. Cortical cells from control root tips. (A) The cell corners lack intercellular spaces. The cell walls (CW) are thin and variable in thickness.
The thinnest regions of cell walls are sinuous (arrowhead). The cytoplasm is dense and contains numerous mitochondria (M), plastids (P) and the
cisternae of ER (circles). The vacuoles (V) are relatively small. The nuclei (N) are centrally located and contain a large nucleolus (NU). The
chromatin is largely uncondensed. TEM. Bar=2 mm. (B) The cells shown here are similar in their ultrastructure to those in (A), except that cell
corners show signs of developing spaces. The cell corners are prominent and contain dense and vacuolate materials (arrowheads). The cytoplasm
is dense and rich in mitochondria (M), plastids (P) and ER (circles). Some plastids contain starch grains. The vacuoles (V) are small, discrete and
also show signs of fusion. N, nucleus. TEM. Bar=2 mm.
Chilling injury in cucumber root 2229
Intermediate stages of chilling injury
Cytoplasmic density was still very high and was
comparable to that in the cells showing signs of
early injuries (Figs 4B, 7A, B). ER cisternae became
more distended and more irregular in their form (Figs
4B, 8A). The cytoplasm also contained other unusual
membraneous formations (Fig. 8A). The nuclear envelope was also markedly distended (Fig. 4B).
Fig. 3. (A) Parts of adjoining cortical cells from a control root tip. The cells are at a slightly more advanced stage of differentiation than those in
Fig. 2B. The intercellular regions (IS) have developed cavities. The cell walls (CW) are thicker and less sinuous. The vacuoles are larger and
show signs of continued enlargement through fusion (arrowheads). The presence of starch in plastids is more common. The mitochondria (M) are
abundant. N, nucleus. TEM. Bar=2 mm. (B) A group of cortical cells from a root tip exposed to cold temperature for 1 h. Several cells show early
signs of chilling injury. The ER is distended. In addition to prominent vacuoles (V) the cytoplasm contains smaller vacuolate structures
(arrowheads). The nuclei (N) are not centrally located and nucleoli (NU) are irregular. The cells marked by unlabelled arrows have a normal
appearance. TEM. Bar=2 mm.
2230 Lee et al.
Mitochondria and plastids became progressively more
swollen and translucent, losing much of their internal
membranes and ribosomes (Figs 4B, 7A, B). Some
mitochondria showed irregular aggregates of remaining/
Fig. 4. (A) Cortical cells from a root tip exposed to cold temperature for 15 min. The cells show signs of both early and intermediate stages of
chilling injury. The cytoplasm is dense and ®lled with numerous distended membraneous structures, most of which are probably ER. The
mitochondria (M) are abundant and their interior appears granular. The plastids (P) appear swollen, contain starch grains, and have a moderately
dense interior. The number and size of vacuoles (V) are highly variable. The nucleus (N) has an irregular shape and contains an irregularly shaped
nucleolus (NU). The cell walls are thin and irregular. TEM. Bar=2 mm. (B) Cortical cells from a root tip exposed to cold temperature for 48 h.
Some cells show intermediate stages of chilling injury. The nucleus (N) is irregular in shape and the nucleolus (NU) is asymmetrically positioned
and corresponds to the nucleus in its shape. The nuclear envelope is grossly distended in places (arrowheads). The ER is also highly distended.
The cytoplasm is highly dense and contains many spherical to oval-shaped bodies, the larger of which may be plastids (P) and the smaller
mitochondria (M). The interior of these bodies appears granular. The plasmalemma has pulled away from the cell wall in places (arrows). The star
indicates a completely damaged cell, which is virtually empty. TEM. Bar=2 mm.
Chilling injury in cucumber root 2231
modi®ed ribosomes (Fig. 7A, B). However, there did
not appear to be a close correlation between the loss in
ribosomes and other changes taking place in mitochon-
dria. For example, there were examples of highly
swollen mitochondria, which had only a few internal
membranes, but contained abundant, apparently normal
Fig. 5. (A) Cortical cells from a root tip exposed to cold temperature for 15 min. Cells show signs of advanced stages of chilling injury. Cell
walls can not be recognized and may have disintegrated. The nuclei (N) are highly irregular and have almost completely lost the integrity of their
envelope. Nucleoli (NU) are still present but are irregular. The interior of the mitochondria (M) is greatly altered and appears translucent. Also,
the envelope appears to have been damaged in some mitochondria. The plastid (P) envelope also appears to be damaged. The ER may have
vesiculated greatly and can not be recognized. The cytoplasmic density is greatly reduced. TEM. Bar=2 mm. (B) Cortical cells from a root tip
exposed to cold temperature for 16 h. The cell marked by an asterisk shows signs of an intermediate stage of chilling injury. The other cells are
greatly transformed and show signs of the ®nal stages of disintegration. The cytoplasm in these cells is sparsely granular and contains scattered
vesicles of varying sizes (arrowheads). The cell walls are extremely thin and show sign of discontinuity (arrow). TEM. Bar=2 mm.
2232 Lee et al.
ribosomes (Fig. 7A). In some instances the remaining
internal membranes in mitochondria had vesiculated
(Fig. 7A). The cell walls were extremely thin and the
plasmalemma had pulled away from the cell wall in
places (Fig. 4B).
Advanced stages of chilling injury
Cytoplasmic density progressively declined (Figs 5A,
8B, 9A, B). In the ®nal stages, the cytoplasm appeared
very thin and electron-lucent, containing sparsely
distributed particles, many of which may be degraded
Fig. 6. A high magni®cation view of cortical cells from control root tips. (A, B) The cytoplasm is dense and contains numerous polysomes.The
ER cisternae are long with a uniform intracisternal spacing between the limiting membranes. The plastids (P) are dense and occasionally contain
starch grains (S). The internal plastid membranes are few. The mitochondria (M) have a moderately dense stroma, which contains ribosomes,
DNA (arrowheads) and numerous infoldings (cristae) of the inner envelope membrane. The mitochondria have a typical double membraneous
envelope; the membranes are parallel with a uniform spacing (circles). N, nucleus; NP, nuclear pore. TEM. Bar=500 nm.
Chilling injury in cucumber root 2233
ribosomes (Fig. 5B). Cell walls were extremely thin,
wavy and disrupted in places (Fig. 5B). In some cases
cell walls had disappeared completely, and thus
partitioning between the nuclei of neighbouring cells
was lost (Fig. 5A). The nuclei had lost the integrity of
their envelope, which appeared highly irregular and
disrupted in places (Fig. 5A). Fragmented membranes
of the nuclear envelope were often seen in proximity
Fig. 7. (A) High magni®cation view of a cortical cell from a root tip exposed to cold temperature for 48 h. The mitochondria (M) are greatly
swollen. They have lost much of their internal membranes, although the envelope is still present. Ribosomes are still in abundance in the
mitochondrial stroma, which also contains some vesiculate, membraneous structures (arrowheads). Membraneous vesicles are also common in the
cytoplasm (arrows), which is highly dense. TEM. Bar=500 nm. (B) A high magni®cation view of a cortical cell from a root tip exposed to cold
temperature for 16 h. The cytoplasm is highly dense. Mitochondria (M) appear swollen. The mitochondrial stroma has lost many of its ribosomes
and thus appears considerably more lucent than the mitochondria of control root tips. A few cristae and the limiting envelope are still present in
some mitochondria. The large vacuolate body (P) with sparse granular inclusions may be a highly swollen plastid. TEM. Bar=500 nm.
2234 Lee et al.
to parent nuclei (Fig. 9A). ER and other cytoplasmic
membranes were highly modi®ed (Fig. 8B) forming
vesicles of varying shapes and sizes (Figs 5A, 9B).
The envelopes of both mitochondria and plastids were
disrupted (Fig. 5A). The cytoplasm also contained
spherical, electron dense bodies (Figs 8B, 9B).
Eventually, the enucleate cell appeared very electron
transparent and sparsely granular, containing vesicles of
varying shapes and sizes (Fig. 5B).
Root pressure measurement
Root pressure in excised cucumber root system measured with the root pressure probe was about 0.15±0.20
MPa when root temperature was kept at 25 °C.
Fig. 8. High magni®cation views of modi®ed cytoplasmic membranes in cortical cells from root tips exposed to cold temperature for 15 min. (A)
The ER is distended. Also present in the cytoplasm are some unusual membraneous formations (arrowheads), TEM. Bar=500 nm. (B) The ER
appears distended and has also vesiculated. The cytoplasm also contains some highly dense deposits (arrowheads). TEM. Bar=500 nm.
Chilling injury in cucumber root 2235
However, when the temperature was gradually lowered
from 25 °C to 8 °C, the root pressure decreased
rapidly, reaching zero MPa within 15±20 min (Fig. 10).
This steep drop in the root pressure is a clear
indication that the cucumber root system is very
sensitive to low root temperature.
Fig. 9. (A) A high magni®cation view of a cortical cell from a root tip exposed to cold temperature for 15 min. The nuclear envelope had broken
down, but the nucleolus (NU) is still present. The membranes (arrowheads) present near the nucleolus may have been part of the nuclear
envelope. TEM. Bar=500 nm. (B) A high magni®cation view of a cortical cell from a root tip exposed to cold temperature for 16 h. The
cytoplasmic density is greatly reduced as compared to the cells in Fig. 7. The mitochondrion (M) present contains only a few cristae and
aggregates of ribosomes and appears highly translucent. Membraneous vesicles (arrowheads) are common in the cytoplasm. The cytoplasm also
contains some dense deposits (arrows). TEM. Bar=500 nm.
2236 Lee et al.
Fig. 10. The effect of low root temperature on cucumber root pressure
(n=6). A steep drop in the root pressure occurred when the
temperature was lowered from 25 °C to 8 °C.
Discussion
It is apparent that some of the cortical cells of cucumber
roots had undergone dramatic ultrastructural changes after
only 15 min of exposure to 8 °C. In addition, all stages of
chilling injury, including those showing disruption and
breakdown of cell walls, nuclei and cytoplasmic
components, were observable after this very short period
of cold exposure. These events are undoubtedly in
response to early changes in the cell components associated with vital cell activities. An early increase in
cytoplasmic density appears to be due to an increase in
the population of free ribosomes, apparently resulting from
the dissociation of polysomes. This means that a loss in the
capacity of cells to synthesize the new proteins that are
required for normal growth and development, occurs very
early on. There are no indications from earlier studies that
the response to cold even among the chilling-sensitive
plants is so rapid, particularly in terms of ultrastructural
modi®cation of cell components leading to cell destruction.
Many of the earlier ultrastructural studies on chilling
stress to plants have been focused on aerial parts; fewer
studies have examined roots. Accumulation of lipids and
dense deposits in response to cold stress has been observed
in various cell components, including plastids, cytoplasm,
regions between the plasmalemma and the cell wall, and in
intercellular spaces (Slack et al., 1974; Ilker et al., 1976;
Platt-Aloia and Thomson, 1976; Wise et al., 1983).
Chloroplasts, mitochondria and cytoplasmic membranes,
including ER, have been observed to undergo dramatic
changes. The swelling of plastids and mitochondria, and
dilation, disorganization and disruption of their membranes, disruption and vesiculation of ER and other
cytoplasmic membranes appear to be common features
among the ultrastructural changes associated with chillinginduced injuries (Millerd et al., 1969; Taylor and Craig,
1971; Forde et al., 1975; Kimball and Salisbury, 1973;
Ilker et al., 1976; Moline, 1976; Niki et al., 1978; Nessler
and Wernsman, 1980; Murphy and Wilson, 1981; Wise
et al., 1983; Ishikawa, 1996). The observations presented
here are consistent with the results of earlier studies and, in
addition, provide substantial new information on these and
other changes related to cytoplasmic, nuclear and cell wall
modi®cations.
Recent biochemical studies on plant response to chilling
stress provide the basis for understanding the rapid
ultrastructural changes observed in cucumber roots in
response to cold exposure. While some studies have shown
no noticeable effect of cold temperatures on mitochondrial
ultrastructure (Nessler and Wernsman, 1980; Wise and
Naylor, 1987), cell membranes and mitochondria generally
appear to be among the main components affected during
cold-induced loss of cell integrity and also during cold
acclimation. It is well known that cold stress affects fatty
acid composition and the ¯uidity of cellular membranes,
proteins and metabolic processes (Graham and Patterson,
1982; Nishida and Murata, 1996). Mitochondria appear to
be one of the main cellular components to be affected (De
Santis et al., 1999), and chilling-acclimation-responsive
nuclear genes, including the catalase gene (Prasad, 1997),
have been isolated. Within a metabolically active plant cell
the involvement of oxygen in the respiratory and
photosynthetic processes leads to the generation of
hydroxyl radicals and other free radicals which can
damage DNA, proteins and lipids (Bowler et al., 1992).
However, plants are equipped with enzymes such as
catalases, peroxidases and superoxide dismutase and free
radical scavengers to protect cell components from
oxidative stress.
The degree of protection against environmental stresses
varies among species, and there are indications that
chilling-sensitive plants either do not produce these
enzymes to the level required for protection from oxidative
stress, or the level of these enzymes goes down in response
to exposure of plants to chilling temperatures. The drastic
ultrastructural changes observed in mitochondria suggest
that the oxidative stress may also be the main cause of this
rapid response in cucumber roots. Monitoring the level of
catalases, peroxidases and superoxide dismutase in cucumber root tips during the periods of cold exposure will be
necessary to con®rm this.
Cold sensing of plants is mediated by an early event in
the signalling process, involving transient in¯ux of
calcium into the cytosol (Knight et al., 1991; Monroy
and Dhindsa, 1995). Oxidative signals also cause a
transient increase in cytosolic calcium (Price et al.,
1994). These studies point to a link between cold sensing,
calcium and oxidative stress, and studies related to
cytochemical localization of calcium in cucumber root
Chilling injury in cucumber root 2237
tips after varying periods of cold exposure are presently
being undertaken by the authors.
The observation of a sudden, steep drop in root pressure
when the temperature of the bathing solution was lowered
to a chilling temperature provides further support for the
high sensitivity of cucumber roots to low temperature. The
challenge now is to understand the biochemical and
structural basis for the rapid response of cucumber to
chilling temperature in its root-water relations.
Acknowledgements
This work was supported by grants from the Korea Science and
Engineering Foundation (KOSEF) to the Agricultural Plant Stress
Research Centre (APSRC, R11-2001-09201004-0) of Chonnam
National University.
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