Journal of Experimental Botany, Vol. 53, No.

372,
Antioxidants and Reactive Oxygen Species in Plants Special Issue,
pp. 13511365, May 2002

Plant responses to abiotic stresses: heavy metal-induced

oxidative stress and protection by mycorrhization
Andres Schu tzendu bel and Andrea Polle1
Forstbotanisches Institut, Abteilung I, Forstbotanik und Baumphysiologie, Georg August Universita t Go ttingen,
Bu sgenweg 2, 37077 Go ttingen, Germany
Received 3 August 2001; Accepted 2 December 2001

Abstract
The aim of this review is to assess the mode of action
and role of antioxidants as protection from heavy
metal stress in roots, mycorrhizal fungi and mycorrhizae. Based on their chemical and physical properties three different molecular mechanisms of heavy
metal toxicity can be distinguished: (a) production of
reactive oxygen species by autoxidation and Fenton
reaction; this reaction is typical for transition metals
such as iron or copper, (b) blocking of essential
functional groups in biomolecules, this reaction
has mainly been reported for non-redox-reactive
heavy metals such as cadmium and mercury, (c) displacement of essential metal ions from biomolecules;
the latter reaction occurs with different kinds of heavy
metals. Transition metals cause oxidative injury in
plant tissue, but a literature survey did not provide
evidence that this stress could be alleviated by
increased levels of antioxidative systems. The reason
may be that transition metals initiate hydroxyl radical
production, which can not be controlled by antioxidants. Exposure of plants to non-redox reactive
metals also resulted in oxidative stress as indicated
by lipid peroxidation, H2O2 accumulation, and an
oxidative burst. Cadmium and some other metals
caused a transient depletion of GSH and an inhibition
of antioxidative enzymes, especially of glutathione
reductase. Assessment of antioxidative capacities
by metabolic modelling suggested that the reported
diminution of antioxidants was sufficient to cause
H2O2 accumulation. The depletion of GSH is apparently a critical step in cadmium sensitivity since
plants with improved capacities for GSH synthesis
displayed higher Cd tolerance. Available data suggest that cadmium, when not detoxified rapidly
enough, may trigger, via the disturbance of the redox
1

control of the cell, a sequence of reactions leading

to growth inhibition, stimulation of secondary metabolism, lignification, and finally cell death. This view
is in contrast to the idea that cadmium results in
unspecific necrosis. Plants in certain mycorrhizal
associations are less sensitive to cadmium stress
than non-mycorrhizal plants. Data about antioxidative
systems in mycorrhizal fungi in pure culture and in
symbiosis are scarce. The present results indicate
that mycorrhization stimulated the phenolic defence
system in the PaxillusPinus mycorrhizal symbiosis.
Cadmium-induced changes in mycorrhizal roots
were absent or smaller than those in non-mycorrhizal
roots. These observations suggest that although
changes in rhizospheric conditions were perceived by
the root part of the symbiosis, the typical Cd-induced
stress responses of phenolics were buffered. It is
not known whether mycorrhization protected roots
from Cd-induced injury by preventing access of
cadmium to sensitive extra- or intracellular sites, or
by excreted or intrinsic metal-chelators, or by other
defence systems. It is possible that mycorrhizal fungi
provide protection via GSH since higher concentrations of this thiol were found in pure cultures of
the fungi than in bare roots. The development of
stress-tolerant plant-mycorrhizal associations may
be a promising new strategy for phytoremediation
and soil amelioration measures.
Key words: Antioxidant systems, heavy metals, mycorrhiza,
oxidative stress.

Introduction
To date an unprecedented, rapid change in environmental
conditions is observed, which is likely to override the

To whom correspondence should be addressed: Fax: [ 49 551 39 2705. E-mail: apolle@gwdg.de

Society for Experimental Biology 2002

1352 Schu tzendu bel and Polle

adaptive potential of plants, especially that of tree species

with their long reproductive cycles. These environmental
changes mainly originate from anthropogenic activities,
which have caused air and soil pollution, acid precipitation, soil degradation, salinity, increasing UV-B radiation, climate change, etc. In addition, plants are exposed
to natural climatic or edaphic stresses, for example, high
irradiation, heat, chilling, late frost, drought, flooding,
and nutrient imbalances. Some of these stress factors
may fluctuate significantly in intensity and duration on
time scales of hours, days, seasons, or years; others
may change slowly and gradually affect plant growth
conditions. Since plants are sessile organisms and have
only limited mechanisms for stress avoidance, they need
flexible means for acclimation to changing environmental
conditions. In order to improve a plants protection, it
is important to understand the mechanisms contributing
to stress tolerance.
A common consequence of most abiotic and biotic
stresses is that they result, at some stage of stress
exposure, in an increased production of reactive oxygen
species (Polle and Rennenberg, 1993). The successive
reduction of molecular oxygen to H2O yields the
intermediates O2Y , HO and H2O2, which are potentially
toxic, because they are relatively reactive compared with
O2. Reactive oxygen species may lead to the unspecific
oxidation of proteins and membrane lipids or may cause
DNA injury. As a consequence, tissues injured by
oxidative stress generally contain increased concentrations of carbonylated proteins and malondialdehyde and
show an increased production of ethylene (Dean et al.,
1993; Ames et al., 1993).
For a long time reactive oxygen species have been
considered mainly as dangerous molecules, whose levels
need to be kept as low as possible. Now this opinion is
changing rapidly. It has been realized that reactive oxygen
species play important roles in the plants defence system
against pathogens (oxidative burst, Alvarez and Lamb,
1997; Doke, 1997; Bolwell et al., 2002), mark certain
developmental stages such as tracheary element formation, lignification and other cross-linking processes in
the cell wall (programmed cell death, Jacobson, 1996;
Teichmann, 2001; Fath et al., 2002) and act as intermediate signalling molecules to regulate the expression of
genes (May et al., 1998; Karpinski et al., 1999; Neill et al.,
2002; Vranova et al., 2002). Because of these multiple
functions of activated oxygen, it is necessary for cells to
control the level of reactive oxygen molecules tightly, but
not to eliminate them completely.
The control of oxidant levels is achieved by antioxidative systems. These defence systems are composed
of metabolites such as ascorbate, glutathione, tocopherol,
etc., and enzymatic scavengers of activated oxygen such
as SODs, peroxidases and catalases (Noctor and Foyer,
1998; Asada, 1999). The maintenance of ascorbate in its
$

reduced form is achieved by monodehydroascorbate

radical reductase (MDAR) and NAD(P)H or ferredoxin
as reductant or by the operation of the ascorbate
glutathione pathway (Foyer and Halliwell, 1976;
Borraccino et al., 1986; Miyake and Asada, 1994). In
the latter pathway the reduction of dehydroascorbate is
coupled to the oxidation of glutathione (GSH), which,
in turn, is reduced by glutathione reductase by oxidation
of NADPH (Foyer and Halliwell, 1976). Antioxidant
systems and their significance for the acclimation of
plants to air pollution and climatic stresses have been
reviewed frequently with emphasis on the responses of
leaves (Smirnoff and Pallanca, 1996; Polle, 1996, 1997,
1998; Smirnoff, 1996; Noctor and Foyer, 1998; Asada,
1999). Less attention has been paid to soil-borne stresses
and their effects in roots.
In soils influenced by human activities a range of
different problems such as overexploitation, salinity,
acidification, and contamination by various pollutants
have been reported. Increasing emissions of heavy metals
are dangerous because they may get into the food chain
with risks for human health (Lantsy and Mackenzie,
1979; Galloway et al., 1982; Angelone and Bini, 1992).
For the recultivation of degraded soils and the reclamation of industrial sites, stress-tolerant plants are required.
Biotechnological efforts are underway to improve plant
stress tolerance and the ability to extract pollutants from
the soil with the aim of using plants for soil clean-up
(Salt et al., 1995). In order to devise new strategies for
phytoremediation and improved tolerance, it is important to understand the basic principles as to how the
pollutants are taken up and act at the cellular and tissue
level. In the present study the occurrence and mode of
action of metal pollutants will be briefly reviewed, and the
role of antioxidants as defence systems will be discussed.
By applying metabolic modelling, oxidant fluxes will be
calculated as an estimate of oxidative stress levels and for
the prediction of efficient compensation mechanisms in
roots. A further question that will be addressed is whether
there is evidence that mycorrhizal symbionts improve
plant performance under heavy metal stress through
increased antioxidative systems.

Occurrence, chemical and physical properties

of heavy metals and their mode of action
Heavy metals are defined as metals with a density higher
than 5 g cm Y 3. 53 of the 90 naturally occurring elements
are heavy metals (Weast, 1984), but not all of them are of
biological importance. Based on their solubility under
physiological conditions, 17 heavy metals may be available for living cells and of importance for organism and
ecosystems (Weast, 1984). Among these metals, Fe, Mo
and Mn are important as micronutrients. Zn, Ni, Cu, V,
Co, W, and Cr are toxic elements with high or low