Documenti di Didattica
Documenti di Professioni
Documenti di Cultura
Submitted
To
DR. M Balakrishnan
Submitted
By
Shashi Meena
Scientist- trainee
FOCARS-101 st
Index
Introduction----------------------------------------------------------------------------------- 3-4
Sources of ROS--------------------------------------------------------------------------------4-6
Types of ROS---------------------------------------------------------------------------------6-10
Superoxide radicals (O2.-)
Singlet oxygen (1O2)
Hydrogen peroxide (H2O2)
Hydroxyl radical (OH.)
Damages due to ROS----------------------------------------------------------------------10-16
Lipid peroxidation
Protein oxidation
DNA damage
Conditions enhancing overproduction of ROS----------------------------------------16-19
Drought
Salinity
Chilling
Metal toxicity
High temperature
ROS scavenging mechanism in plants------------------------------------------------19-20
Non-enzymatic antioxidants----------------------------------------------------------20-28
Ascorbic acid (Vitamin-C)
Glutathione (GSH)
Tocopherol (Vitamins-E)
Carotenoid
Phenolic compounds
Proline
Enzymatic antioxidants----------------------------------------------------------------28-33
Superoxide dismutase (SOD)
Catalase (CAT)
Guaiacol peroxidase (GPOX)
Glutathione reductase (GR)
Monodehydroascorbate reductase (MDHAR)
Dehydroascorbate reductase (DHAR)
Ascorbate peroxidase (APX)
Glutathione-S-transferase (GST)
Glutathione peroxidase (GPX)
Conclusion------------------------------------------------------------------------------------34
References-----------------------------------------------------------------------------------35-37
burst is the result of ROS production either extracellular or intracellular, which has negative
impact on physiological and biochemical functions of the plants (Gill et al., 2010). Plants
must be able to sense the environmental cues through sensors before being able to respond
appropriately to the abiotic stress. A signal transduction pathway invoked after initial
recognition of stress which ultimately result in the production of secondary messenger,
involved in the activation of stress induced genes. Under normal conditions ROS act as
signalling molecule but under stress condition it results in oxidative stress or burst in cell.
Sources of ROS:CHLOROPLA
ST
MITOCHONDR
IA
PLASMA
MEMBRANE
ROS
CELL
WALL
APLOPLAS
T
PEROXISOM
E
ENDOPLASMIC
RETICULUM
GLYOXYSOM
E
catalysed by enzyme lipoxygenase (LOX) (Asada, 2006; Foyer et al., 2009). Due to stress
conditions, there is limited CO2 fixation by Calvin cycle leads to a decrease in oxidized
NADP+ which serves as electron acceptor in photosynthesis. Stress conditions lead to
overloading of the ETC, result in leakage of electron from PSI (Photosystem I) to O 2,
reducing it to O2.-. This process is known as Mehler reaction (Karunppanapandian et al.,
2011). One of the major problems that plants encounter during stress is the uncoupling of the
light reaction of photosynthesis from the fixation of CO2 and production of highly reactive
chlorophyll molecules which ultimately result in the production of highly reactive singlet
oxygen species.
ROS production in peroxisome is the result of their essentially oxidative pathways and is the
major site of intracellular ROS production. Peroxisomes also produce superoxide radicals
through normal metabolism. Glycolate oxidase is the enzyme of photorespiratory pathway
(C2) in plants which involved in the oxidation of glycolate to glyoxylate and also result in the
production of H2O2 (Mittler et al., 2004). There are two sites of O2.-generation in peroxisomes
(del Rio et al., 2002). One site is located in the organelle matrix, in which enzyme xanthine
oxidase (XOD) is present and another site is located in the peroxisomal membranes which
have NAD(P)H-dependant ETC. Xanthine oxidase catalyzes the oxidation of xanthine and
hypoxanthine to uric acid and a well-known producer of superoxide radicals (Lopex-huertus
et al., 1999; del Rio et al., 2002). The apoplast is alsoan important site for H2O2 production
in plants under adverse environmental conditions such as drought and salinity (Zhu et al.,
2001). The two enzymes are involved in apoplastic ROS production such as oxalate oxidase
and amine oxidase (Mittler, 2002), catalyse the oxalic acid and causes oxidative deamination
of ployamines (i.e. putrescine, spermine, and spermidine) respectively (Cona et al., 2006).
The various electron transporting oxidoreductases such as NADPH oxidase and quinine
reductase are present at plasma membranes and lead to production of free radicals at plasma
membrane under stress conditions (Heyno et al., 2011).
ROS production in mitochondria takes place under normal respiratory conditions but can be
enhanced in response to various environmental stress conditions. The primary source of
mitochondrial ROS generation is the mitochondrial ETC, which harbours electrons with
sufficient energy to directly reduce O2 (Rhoades et al., 2006). The secondary source of ROS
production in mitochondria is mitochondrial matrix which contains several enzymes. Cell
walls are the major sites for active metabolism and ROS production in plants. NADH oxidase
localised to the plasmalemma and involved in defence mechanism against pathogens and
pests. NADH oxidase give rise to H2O2 catalyzing NADH, generated by cell wall associated
malate dehydrogenase enzymes (Higuchi, 2006). Diamine oxidase, Lipoxygenase and cell
wall localised peroxidases are also involved in the production of activated O 2 during stress
conditions. The endoplasmic reticulum is also involved in superoxide radical generation
through NAD(P)H- dependent electron transport involving cytochrome P450 (Mittler, 2002).
Types of ROS:Chloroplast, mitochondria or peroxisomes are the organelles with highly oxidizing metabolic
activity or with intense rate of electron flow. These two activities are major sources of ROS in
plant cells (Gill et al., 2010). Plants have ability to use O2 for the energy production and this
energy is utilized for their own developmental processes. It has been estimated that 1-2% of
O2 consumed by plants is contributed to ROS production in various subcellular loci (Blokhina
et al., 2003). ROS is derived from ground state oxygen either by energy transfer or by
electron leakage reactions. The former reaction leads to the formation of singlet oxygen ( 1O2),
whereas the latter reaction results in the sequential reduction to superoxide (O 2.-), hydrogen
peroxide and hydroxyl radical (Klotz, 2002). In plants active oxygen species are continuously
6
Superoxide radical
ion (O2.-)
Perhydroxyl
radical
(HO2.)
e-
O23-
Oxene
ion (O-)
Water
(H2O)
Hydroxy
l radical
(OH.)
O2-
Water
carried out either spontaneously or by superoxide dismutase (SOD). Furthermore, O 2.can also donate electron to Fe3+ to yield a reduced form of iron (Fe 2+). This reduced
form of iron (Fe2+) oxidized by H2O2 and rapidly generate OH. . This reaction is the
Fentons reaction or the Haber-Weiss reaction.
The generations of superoxide radicals also trigger the production of more reactive
ROS like OH. and 1O2. These reactive oxygen species may cause membranes lipid
peroxidation and cellular weakning. The major site of superoxide radical formation is
the thylakoid membrane bound primary electron acceptor of PSI during
photosynthesis. It is also produced during aerobic respiration (Arora et al.,2002).
1. Singlet oxygen:-1O2 is an unusual form of ROS in plants. It can be produced by
insufficient energy transfer during photosynthesis to chlorophyll, result in the
formation of triplet state chlorophyll (3Chl*). This triplet state can react with dioxygen
(3O2) to give up the very reactive 1O2. For the formation of 1O2, electron transfer to O2
does not require. The life span of singlet oxygen within the cell is approximately 3s
(Hatz et al., 2007; Hackbarth et al., 2010). It has been found that a fraction of 1O2 has
been able to diffuse over considerable distances of several 100 nanometers (nm). It
has been also found that 1O2 can last for 4s in water and 100s in a non-polar
environment (Foyer et al., 1994). 1O2 reacts with most of the biological molecules and
it directly oxidizes unsaturated fatty acid, protein and DNA (Foyer et al., 1994). It is
documented to be the most reactive species responsible for light-induced loss of
Photosystem II (PSII) activity which may trigger cell death (Liszkay et al., 2008). It is
well established that if 1O2 is produced through normal metabolic pathways, it is
efficiently quenched by -carotene, tocopherol or plastoquinone. If it is not quenched
by these molecules, then it can activate the upregulation of genes, involved in the
molecular defense responses against photo-oxidative stress (Liszkay et al., 2008;
Lizskay, 2005).
1997). OH. is also responsible for the oxidation of organic substrates, may proceed by
two possible reactions either by addition of OH. to an organic molecule or by
abstraction of a hydrogen atom from it. These two reactions which involve oxidation
of organic substrates, also leads to the production of destructive free radicals such as
alkoxy, peroxy, semiquinones, reduced H2O2 etc., besides toxic OH. (Arora et al.,
2002).
Damages due ROS:In order to avoid oxidative damages or stress, the production and removal of ROS must be
strictly regulated. Under various stressful conditions such as drought, salinity, high light,
metal toxicity, temperature and uv-radiation, the balance between production and scavenging
of ROS is perturbed. Oxidative stress is the condition in which the level of ROS exceeds
the level of antioxidants in the cell. High level of ROS can cause damage to various
biomolecules such as lipids, DNA and proteins. ROS not only affect these molecules, it can
also alter intrinsic membrane properties like ion transport, fluidity, inactivation of enzyme,
cross-linking of protein, inhibition of protein synthesis, damage to DNA, etc. ultimately
resulting in autophagy (cell death).
Fig: The graph showing relationship between
ANTIOXIDANT
S
STRESS
ABIOTIC
CONDITIONSSTRESS
ROS
Protein
Oxidative
damage 10
Lipid
peroxidation
Enzyme
inactivation
Alter membrane
properties
DNA
damage
Damage to
photosynthetic
machinery
CELL
DEATH
Initiation:- This steps also involves transition metal complex, especially those of Fe
and Cu. O2.- when react with H2O2 give rise to OH., highly reactive. This free radical
initiates the lipid peroxidation in a membrane by reacting with a polyunsaturated fatty
acid (PUFA). This reactions result in the formation of lipid alkyl radical (R.). This
further converted into ROO.- (lipid peroxy radical) in aerobic condition. A single
initiation event has the potential to generate multiple peroxide molecules by a chain
reaction (Gill et al., 2010).
11
R. + H2O
(Lipid)
Propagation:- The peroxy radical formed is highly reactive and able to propagate the
chain reaction.
Propagation steps:R. + O2
ROO.
(Lipid peroxy radical)
ROO. + RH
ROOH + R.
RO.Epoxides, hydroperoxides, glycol, aldehydes
ROOH
Termination:- The LPO reaction stopped when two free radicals combine together
and form fatty acid dimer.
Termination steps:R. + R.
R+R
(Fatty acid dimer)
R. + ROO.
ROOR
(Peroxide bridged dimer)
ROO. + ROO.
ROOR + O2
(Peroxide bridged dimer)
Lipid peroxidation events decreases membrane fluidity, make it leaky, damage membrane
proteins, inactivation receptors, enzymes, ion channels (Moller et al., 2007). PUFA
12
peroxidation result in the production of MDA and HNE (2-hydoxy-2-nonenal), from linolenic
and linoleic acids respectively.
Protein oxidation:-ROS production under biotic and abiotic stresses, does not only
initiates LPO but also result in covalent modification of proteins. Protein
conformation and stability is dramatically affected by sudden changes in the
environment, giving rise to protein unfolding, misfolding and aggregation. Folded
states represent the most stable forms under native conditions, but partially folded
states that allow for efficient interaction with binding partners are of fundamental
importance in biological activity. Protein function is dependent on its unique threedimensional structure that is adopted by the initial folding of the polypeptide chains
after translation. Folded proteins are generally much less prone to aggregation and
13
14
can be damaged in oxidative conditions by reacting with LPO products, such as HNE
and MDA.
DNA damage:- DNA is the genetic material of the cell and any damage to the DNA
can result in malfunctions or complete inactivation of encoded proteins. DNA can be
damaged by ROS production under biotic and abiotic stresses. High level of ROS not
only damage to cell structures, lipids and proteins but also damage nucleic acid (Valko
et al., 2006). ROS can cause oxidative damages to nuclear, mitochondrial, and
chloroplastic DNA. Nuclear DNA is more resistant to oxidative damage than
mitochondrial and chloroplastic DNA due to the presence of protective proteins and
histones (Richter, 1992). High level of ROS or oxidative stress attack on DNA results
in deoxyribose oxidation, removal of nucleotides, strand breakage, modification of
bases and DNA protein crosslinks, also enhance the chances of mutations (causes
G:C alterations).ROS attacked on both sugar and base moieties of DNA. ROS attack
on DNA bases generally involves OH .addition to double bond, while sugar damage
mainly involves hydrogen abstraction from the deoxyribose. It has been reported that
OH. is most reactive radical and causes damage to purine and pyrimidine bases and
also deoxyribose backbone (Gill et al., 2010). Upon reaction with DNA, it results in
the production of C-8 hydroxylation of quinine to form 8-oxo-7,8 dehydro-2deoxyguanosine etc (Tsuboi et al., 1998). 8-hydroxyguanine is the most commonly
observed product during oxidative stress. Singlet oxygen is dangerous to guanine
only, whereas H2O2 and O2.- do not react with bases at all. ROS can indirectly attack
DNA bases by producing reactive molecules from lipid peroxidation. When the plants
are exposed to various environmental stresses such as salinity (Liuet al., 2000) and
metal toxicity (Meriga et al., 2004) enhances DNA degradation. ROS attack to DNA
sugars result in single-strands breaks. It has been reported that DNA protein cross
15
links formed when OH. attacks on either on DNA or protein associated with it. DNA
damage affects various physiological functions of plants (Gill et al., 2010).
Conditions enhancing overproduction of ROS:Under normal growth conditions, the production of ROS in plants is low. However, in
response to various stress conditions, the generation of toxic O2 species is drastically
enhanced in plants disturbing the balance between production and scavenging of O2.-,
OH. and H2O2 in the intracellular environment (Sharma et al., 2010). The effects of
various stress factors such as drought, salinity, chilling, metal toxicity, ROS
production are described below:1. Drought:- ROS production in plant is enhanced under drought stress by several
ways. Several activities under drought stress results in overproduction of ROS
through the chloroplast Mehler reaction (Asada, 1999). During drought stress, there
is inhibition of CO2 assimilation which in turn, leads to reduced NADP+ generation
through the C3 cycle. This result in lack of NADP+ electron acceptor, over reduction
of the photosynthetic ETC occurs which leads to a higher leakage of electrons to O 2
by the Mehler reaction (Sharma et al., 2012). Under drought stress, there is an
imbalance between light capture and its utilization, this result in decreased in
photosynthetic activity in plant tissues (Foyer et al., 2000). The other reason of
ROS generation is dissipation of excess light energy in the PSII core and antenna,
which are potentiallly dangerous under drought stress. Under drought stress, the
photorespiratory pathway is also enhanced and there is more RUBP oxygenation
due to limitation in CO 2 assimilation (Noctor et al., 2002). Sangmein lee and
Chung-MO Park found that the NTL4 gene is induced in the response of H 2O2, and
also related with the accumulation of ROS under drought stress. Under drought
stress conditions, 70% of total H2O2 production is contributed by photorespiration
16
17
to reduction in growth and productivity of crop plants (Fryer et al., 1998; Prasad,
1997; Zhang et al., 2008).
4. Metal toxicity:- Metals are essential for functioning of physiological and
biochemical processes and consequently for normal growth and development of
plants. The increasing level of metals into the environment has negative impact on
plant growth and metabolism, ultimately leading to reduce crop yields (Salt et al.,
1995; Mishra and Dubey, 2005). Net photosynthesis decreases due to damage to
photosynthetic metabolism, including photosynthetic electron transport under metal
stress conditions. Alter photosynthetic metabolism lead to overproduction of ROS.
Metal stress affects various mechanisms: such as interference with functional sites
in proteins, displacement of essential elements, thereby disturbing enzymatic
functions, enhanced ROS production. Heavy metal stress causes various problems
in plants:- Induction of peroxidation which can directly cause biomembrane
deterioration, decomposition of polysaturated fatty acids (PUFAs) result in the
production of oxidative stress (Karunppanapandian and Manoharan, 2008;
Karunppanapandian et al., 2009, 2011).
5. High temperature:- Temperature stress is one of the major type of abiotic stress
and has devastating effects on plant growth and metabolism. High temperature is
the result of global climate change and a critical factor for plant growth and
productivity; high temperature is now considered as major stresses for restricting
crop production. The growth and development of plants involves a large number of
biochemical reactions, all of which are sensitive to some degree to temperature.
Consequently, plant responses to high temperature vary with the extent of the
temperature increase, its duration, and the plant type. When plants exposed to high
temperature stress leads to excess accumulation of toxic compounds, especially
reactive oxygen species (ROS). In response to high temperature, the reaction
18
(31.9C/27.8C)
decreased
chlorophyll
(Chl)
content
and
peroxidase
(GPOX),
19
Superoxide
dismutase
(SOD),
Monodehydroascorbate
redutase
(MDHAR),
Dehydroascorbate
reductase
Glutathione-S-transferases (GST),
20
(Smirnoff, 2000) by directly reacting with O2.-, H2O2 and by preserving the
enzyme activities which contain prosthetic transition metal ions (Noctor and
Foyer, 1998). H2O2 is removed from the cells through Ascorbate-Glutathione
cycle (Pinto et al.,2003). The major pool of ascorbate is contributed by
Smirnoff- Wheelar pathway(Smirnoff, 2000).
Oxidation of ascorbic acid occurs in two sequential steps:- By producing
monodehydroascorbate (MDHA) either it may re-reduced to ascorbate or it
may disproportionates into ascorbic acid and dehydroascorbate (DHA). In the
Asada-Halliwell pathway (H2O2 scavenging pathway), two molecules of
ascorbic acid are utilized by ascorbate peroxidase to reduce H 2O2 to water and
MDHA. MDHA if not rapidly converted into ascorbate, it can spontaneously
dismutate into DHA and ascorbate and it is reduced to ascorbate by
Monodehydroascorbate reductase which is NAD(P)H-dependent enzyme.
DHA is highly unstable at pH values greater than 6.0 and is decomposed to
tartarate and oxalate (Noctor and Foyer, 1998). DHA is also contributed to
ascorbic acid pool by the enzyme dehydroascorbate reductase which utilized
reducing equivalents from Glutathione (Asada, 1996).
The biosynthesis of ascorbate within the chloroplast provides a putative
mechanism for the regulation of electron transport. Ascorbate is not only a
potent antioxidant, but it is also involved in pH-meditated modulation of PS II
activity and its down regulation is associated with Zeaxanthin formation
(Neubauer and Yamamoto, 1992). This is a potent mechanism for preventing
photo-oxidation. It has been estimated that the level of Ascorbic acid is alter in
various biotic and abiotic stresses ( Mishra et al., 2011). The level of ascorbic
acid is enhanced by overexpressing the enzymes involved in ascorbic acid
biosynthesis confers abiotic stress tolerance (Chaves et al., 2002). But,it has
21
also been observed that in the roots and nodules of Glycine max under Cd
stress result in decrease in the ascorbate. Cd stress also decrease the Ascorbate
in Cucumin sativuschloroplast and in the leaves of A. thalianaand P.sativum(Li
et al., 2010).
2. Glutathione (GSH):- GSH is an abundant tripeptide (-glutamyl cystein
glycine) of low molecular weight non-protein thiol in plant tissues that plays
an important role in intracellular defense against oxidative damage induced by
ROS. In plant tissues it occurs abundantly in reduced form. It is localized in all
cell compartments like cytosol, vacuole, chloroplast, endoplasmic reticulum,
peroxisome, mitochondria as well as in apoplast (Mittler et al, 1992), here it
perform several physiological functions like detoxification of xenobiotics,
sulfate transport regulation, signal transduction and regulate the expression of
stress responsive genes. It also plays an important role in diverse biological
events including cell division, cell differentiation, cell death and senescence,
pathogen resistence, synthesis of proteins and nucleic acids, synthesis of
phytochelatins for metal chelation and enzymatic regulation (Foyer et al.,
1997).
Dehydroascorb
ate reductase
Glutathione
reductase
22
23
25
5. Phenolic compounds:- Phenols also protect plants form toxic ROS. These are
diverse
secondary
metabolites
including
flavonoids,
tannins,
flavones,
isoflavones
and
anthocyanin.
Under
adverse
decreased degradation of proline. It provide protection against ROS inducedoxidative damage by acting as an osmoprotectant, a metal chelator, a protein
stabilizer, an inhibitor of lipid peroxidation, and OH . and 1O2 scavenger
(Trovato et al., 2008). Proline is more effective scavenger of ROS mainly OH .
than sorbitol, mannitol, myo-inositol. Proline plays an important role in
maintaining the level of NADPH in cells by potentiating pentose-phosphate
pathway. Through this Proline is able to maintain GSH and ASH in the
reduced state. Proline also accumulate in water stress deficit conditions
(Stewart, 1981) and perform an important function as protective compatible
osmolyte in scavenging free radicals and facilitates a correction of altered
redox potential (Harre et al., 1999).
Enzymatic antioxidants:1. Superoxide dismutase (SOD):- SOD is a metalloenzyme and the most effective
intracellular enzymatic antioxidant that plays central role in defense against oxidative
stress induced by ROS in all aerobic organisms (Gill et al., 2010). This enzyme is
present in most of the subcellular compartments that generate activated oxygen (Chen
et al., 2010). This enzyme removes superoxide radical by dismutation into O 2 and
H2O2 and hence decreases the risk of OH . formation (more toxic than H2O2) through
the metal catalyzed Haber-Weiss reaction. SODs are classified into three main types
depending on their metal cofactors: Cu/Zn SOD is structurally somewhat different
from other two SODs, this isozyme is localized to chloroplasts, peroxisomes and
cytosol; Fe-SOD is localized to chloroplasts and Mn-SOD is localized to
mitochondria (del Rio et al., 1996). Different forms of SOD are nuclear encoded and
by an amino terminal targeting sequence, able to targeted to their respective
subcellular compartment.
27
There are two different techniques to assess the activity of SOD isozymes: negative
staining and to check sensitivity towards KCN and H 2O2. The Cu/Zn-SOD is sensitive
to both inbibitors but Fe-SOD is resistant to KCN and sensitive to H 2O2 (del Rio et
al., 1998). The prokaryotic Mn-SOD and Fe-SOD, and eukaryotic Cu/Zn-SOD
enzymes are dimers, whereas Mn-SOD of mitochondria is tetramer. There is
upregulation of SODs in combating oxidative damages caused due to various stresses
and have a central role in providing tolerance under stress environment. In Mulberry
plant, there is significant increase in SOD activity under saline condition (Harinasut,
2003). In Arabidopsis, it has been reported that there are three genes for Fe-SOD and
Cu-SOD and one gene for Mn-SOD (Kliebenstein, 1999). The activity of SOD is
directly proportional to the increase tolerance of the plant against environmental
stresses.
2. Catalase:- Catalase is a tetrameric heme-containing antioxidant enzyme. It was the
first enzyme to be discovered and characterized. This enzyme is ubiquitous in nature
and its function is to catalyze the dismutation of two molecules of H 2O2 to water and
O2/minute, thus it has highest turnover rates (Gill et al., 2010) but a much lower
affinity for H2O2 than ascorbte peroxidase (APX). In plants, peroxisome is the major
site for H2O2 generation during photorespiratory oxidant, -oxidation of fatty acids,
purine catabolism, photorespiration and other enzymes systems such as XOD coupled
to SOD (del Rio et al., 2006). Catalase shows strong activity towards H2O2 but have
weak activity against organic peroxides. Catalase does not require reducing
28
It catalyzes the regeneration of ascorbic acid from MDHA using NAD(P)H as the
electron donor (Hossain et al., 1985). MDHAR is the only enzyme which use organic
radical as a substrate (MDHA) and also able to reduce phenoxy radicals which are
generated by horseradish peroxidase with H2O2 (Sakihama et al., 2000). This enzyme
is widespread in plants. It also involves in the photoreduction of dioxygen to O 2.
when the substrate MDHA is absent (Miyake et al., 1998). It has been noted that the
overexpression of MDHAR in transgenic tobacco increased tolerance against salt and
osmotic stresses (Eltayeb et al., 2007).
6. Dehydroascorbate reductase (DHAR):- DHAR is the second enzyme involved in
the regeneration of reduced ascorbate from oxidized state (DHA) using GSH as the
reducing substrate (Ushimaru et al., 1997). Reduced ascorbate is major antioxidant in
plants that detoxify ROS, provide tolerance and maintains photosynthetic function
under various stress conditions. DHAR is an important regulator of ascorbic acid
recycling. Overexpression of DHAR activity results in increased tolerance to various
stresses.
7. Ascorbate peroxidase (APX):- This enzyme is involved in the scavenging of ROS
(H2O2) from chloroplasts and cytosol of plant cells. It is a central component of ASHGSH cycles and also involved in H2O2 scavenging in water-water cycle, and plays an
essential role in the control of intracellular ROS levels. This enzyme utilizes two
molecules of ascorbate as hydrogen donor to reduce H 2O2 to water and production of
two molecules of MDHA. The APX family consists of at least 5 different isoforms
including thylakoid, glyoxisome, membrane as well as chloroplast stromal and
cytosolic form (Noctor and Foyer, 1998). The cytosolic form of APX are two in
number which have several functions like H2O2
31
et al., 2005). It is more efficient in H2O2 scavenging than catalase and peroxidase and
it may have a key role in the maintained ROS level during stress. It is a member of
class I super family of heme peroxidase (Welinder, 1992) and regulated by redox
signals and H2O2 (Patterson et al.,1995). Many coworkers have reported enhanced
activity of APX in response to various stresses.
8. Glutathione-S-transferases (GST):- It catalyses the conjugated product of
electrophilic xenobiotic substrates with the tripeptide glutathione (GSH: -glu-cysgly). The plant glutathione transferases, formerly known as glutathione-S-tansferases.
It plays a major role in the detoxification of herbicides and hydroxyperoxide,
hormone sequestration, metabolism of tyrosine, anthocyanin sequestration in vacuole,
apoptosis regulation and in plant responses to various biotic and abiotic stresses
(Dixon et al., 2010). It protects DNA, RNA and proteins from oxidative damages by
removing cytotoxic or genotoxic compounds. GST with the help of GSH can also
reduce peroxides and produce scavengers of cytotoxic and genotoxic compounds.
This enzyme generally found in cytoplasm but various isoforms has also been
reported from microsomal, plastidic, nuclear and apoplastic (Frova, 2003). It has also
been found that GST overexpression also enhance plant tolerance to various abiotic
stresses.
9. Glutathione peroxidase (GPX):- GPX use tripeptide glutathione (GSH) to reduce
H2O2, organic and lipid hydroperoxides, and therefore defense plant cells from
oxidative stress (Noctor et al., 2002).
Conclusion:-
32
ROS are produced in both stressed and non-stressed conditions in plants; however, under
adverse conditions, the equilibrium between production and their scavenging is disturbed.
When the level of ROS exceeds the antioxidants, a cell is said to be in a state of Oxidative
stress. Under favourable growth condition, ROS production is low in different cellular
compartment which is produced as by-products of various metabolic pathways. Various biotic
and abiotic stresses lead to the enhanced production of ROS in plants which are destructive in
nature and cause degradation of pigments, carbohydrates, proteins, lipids, nucleic acids,
inactivation of enzymes, destruction of membranes and vital cellular organelles in plants
which ultimately result in cell death. Under stress conditions, several forms of ROS are
produced such as free radical (superoxide radical; hydroxyl radical; perhydroxy radical and
alkoxy radical) and molecular forms include hydrogen peroxide and singlet oxygen. In
addition to destructive macromolecules at high concentrations, ROS also act as a diffusible
signalling molecule at low concentration that perform role in signal transduction pathways as
well as act as a secondary messenger in various developmental pathways and mediate several
plant responses under stress conditions. To mitigate the harmful effects of ROS, plants
possesscomplex antioxidative defence systems. These defence systems comprising both
enzymatic and non-enzymatic components which are involved in the efficient scavenging of
excess ROS produced during various stresses conditions.
References:1.
2.
A. Arora, R. K. Sairam and G.C. Srivastava, Oxidative stress and antioxidative system in plants,
Current Science, vol. 82, no. 10, pp. 1227-1238, 2002.
A. Cona, G. Rea, R. Angelini, R. Federico, and P. Tavladoraki, Functions of amine oxidases in plant
development and defence, Trends in Plant Science, vol. 11, no. 2, pp. 8088, 2006.
33
3.
4.
5.
6.
7.
8.
9.
10.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
22.
23.
24.
25.
B. Halliwell, Reactive species and antioxidants. Redox biology is a fundamental theme of aerobic
life, Plant Physiology, vol. 141, no. 2, pp. 312322, 2006.
B. Meriga, B. K. Reddy, K. R. Rao, L. A. Reddy, and P. B. K. Kishor, Aluminium-induced production
of oxygen radicals, lipid peroxidation and DNA damage in seedlings of rice (Oryza sativa), Journal
of Plant Physiology, vol. 161, no. 1, pp. 6368, 2004.
C. H. Foyer and G. Noctor, Redox regulation in photosynthetic organisms: signaling, acclimation,
and practical implications, Antioxid Redox Signal,vol. 11:pp. 861905, 2009.
C. H. Foyer and G. Noctor, Oxygen processing in photosynthesis: regulation and signalling, New
Phytologist, vol. 146, no. 3, pp. 359388, 2000.
C. H. Pang and B. S. Wang, Oxidative stress and salt tolerance in plants, in Progress in Botany, U.
Luttge, W. Beyschlag, and J.Murata, Eds., pp. 231245, Springer, Berlin, Germany, 2008.
C. Xiang, B.L. Werner, E.M. Christensen, D.J. Oliver, The biological functions of glutathione
revisited in Arabidopsis transgenic plants with altered glutathione levels, Plant Physiol., vol. 126, pp.
564-574, 2001.
E. F. Elstner, Mechanisms of oxygen activation in different compartments of plant cells, in Active
Oxygen/Oxidative Stress and Plant Metabolism, E. J. Pell and K. L. Steffen, Eds., pp. 1325,
American Society of Plant Physiologists, Rockville, Md, USA, 1991.
E. Lopez-Huertas, F. J. Corpas, L. M. Sandalio, L. A. del Ro, Characterization of membrane
polypeptides from pea leaf peroxisomes involved in superoxide radical generation, Journal of
Biochemistry, vol. 337, pp. 531536, 1999.
Fridovich, I., Annu. Rev. Biochem., vol. 65 pp.97-112, 1995.
G. Noctor and C. H. Foyer, Ascorbate and glutathione: keeping active oxygen under control, Annual
Review of Plant Biology, vol. 49, pp. 249279, 1998.
G. Noctor, S. Veljovic-Jovanovic, S. Driscoll, L. Novitskaya,C. H. Foyer, Drought and oxidative
load in the leaves of C3 plants: a predominant role for photorespiration? Ann. Bot., vol. 89, pp. 841
850, 2002.
G. Noctor, L. Gomez, H. Vanacker, C.H. Foyer, Interactions between biosynthesis,
compartmentation, and transport in the control of glutathione homeostasis and signalling, J. Exp.
Bot., vol. 53, pp. 1283-1304, 2002.
I.M. Moller, P.E. Jensen, A. Hansson, Oxidative modifications to cellular components in plants,
Annu. Rev. Plant Biol., vol. 58, pp. 459-481, 2007.
J. A. Hernndez, M. A. Ferrer, A. Jimnez,A. R. Barcel, F. Sevilla, Antioxidant systems and
O2/H2O2 production in the apoplast of pea leaves. Its relation with salt-induced necrotic lesions in
minor veins, Plant Physiol., vol. 127, pp. 817831, 2001.
K. Asada, Production and scavenging of reactive oxygen species in chloroplasts and their functions,
Plant Physiology, vol. 141, no. 2, pp. 391396, 2006.
K. Asada, Radical production and scavenging in the chloroplasts, in Photosynthesis and the
Environment, N. R. Baker, Ed., pp. 123150, Kluwer, Dordrecht, The Netherlands, 1996.
K. Asada, The water-water cycle in chloroplasts: scavenging of active oxygens and dissipation of
excess photons, Annual Review of Plant Biology, vol. 50, pp. 601639, 1999.
K. J. Davies, Protein damage and degradation by oxygen radicals. I. general aspects, Journal of
Biological Chemistry, vol. 262, no. 20, pp. 98959901, 1987
L.A. del Ro, F.J. Corpas, L.M. Sandalio, J.M. Palma, M. Gmez, J.B. Barroso, Reactive oxygen
species, antioxidant systems and nitric oxide in peroxisomes, J. Exp. Bot., vol. 53, pp. 1255-1272,
2002.
L-O Klotz, Oxidant-induced signaling: effects of peroxynitrite and singlet oxygen, Biol. Chem.,
vol.383, pp. 44356, 2002.
M. Valko, C.J. Rhodes, J. Moncol, M. Izakovic, M. Mazur, Free radicals, metals and antioxidants in
oxidative stress-induced cancer, Chem. Biol. Interac., vol. 160, pp. 1-40, 2006.
M. A. Torres, J.L. Dangl, J. D. G. Jones, Arabidopsis gp91phox homologues AtrbohD and AtrbohF
are required for accumulation of reactive oxygen intermediates in the plant defense response, Proc.
Natl. Acad . Sci ., USA , vol. 99, pp. 517522 , 2002.
N. Navrot, N. Rouhier, E. Gelhaye, J.P. Jaquot, Reactive oxygen species generation and antioxidant
systems in plant mitochondria, Plant Physiology, vol. 129, pp. 185-195, 2007.
34
26. N. Smirnoff, Ascorbate, tocopherol and carotenoids: metabolism, pathway engineering and functions.
in: N. Smirnoff (Ed.), Antioxidants and Reactive Oxygen Species in Plants, Blackwell Publishing
Ltd., Oxford, UK, pp. 53-86, 2005.
27. P. Sharma, A. Bhushan Jha, R. Shankar Dubey, and M. Pessarakli, Reactive oxygen species, oxidative
damage, and antioxidative mechanisms in plants under stressful conditions Journal of Botany, pp. 126, 2012.
28. P. Sharma, A. B. Jha, and R. S. Dubey, Oxidative stress and antioxidative defense system in plants
growing under abiotic Stresses, in Handbook of Plant and Crop Stress,M. Pessarakli, Ed., pp. 89
138, CRC Press, Taylor and Francis Publishing Company, Fla, USA, 3rd edition, 2010.
29. P. Ghezzi, V. Bonetto, Redox proteomics: identification of oxidatively modified proteins,
Proteomics, vol. 3, pp. 1145-1153, 2003.
30. Prasad PVVPisipati SR, Momilovi I, Ristic Z. Independent and combined effects of high
temperature and drought stress during grain filling on plant yield and chloroplast EF-Tu Expression in
spring wheat,.Journal of Agronomy Crop Science, vol. 197, pp.430-441, 2011
31. R. Mittler, Oxidative stress, antioxidants and stress tolerance, Trends Plant Sci., vol. 7, pp. 405-410,
2002.
32. R. Mittler , S. Vanderauwera, M. Gollery, F. Van Breusegem, Reactive oxygen gene network of
plants, Trends Plant Sci., vol. 9, pp. 490498, 2004.
33. S. Bhattacharjee, The language of reactive oxygen species signaling in plants, Journal of Botany,
Vol.no.pp. 1-22, 2012.
34. S. Bhattachrjee, Reactive oxygen species and oxidative burst: roles in stress, senescence and signal
transduction in plant,Current Sci., vol. 89, pp. 1113-1121, 2005.
35. S. Singh Gill and N. Tuteja, Reactive oxygen species and antioxidant machinery in abiotic stress
tolerance in crop plants, Plant Physiology and Biochemistry, vol. 48, pp. 909-930, 2010.
36. S. Kukreja, A.S. Nandval, N. Kumar, S.K. Sharma, S.K. Sharma, V. Unvi, P.K. Sharma, Plant water
status, H2O2 scavenging enzymes, ethylene evolution and membrane integrity of Cicer arientinum
roots as affected by salinity, Biol. Plant., vol. 49, pp. 305-308,2005.
37. S. Hatz, J. D. C. Lambert, and P. R. Ogilby, Measuring the lifetime of singlet oxygen in a single cell:
addressing the issue of cell viability, Photochemical and Photobiological Sciences, vol. 6, no. 10, pp.
11061116, 2007.
38. S. Hackbarth, J. Schlothauer, A. Preu, and B. Roder, New insights to primary photodynamic effects
singlet oxygen kinetics in living cells, Journal of Photochemistry and Photobiology B, vol. 98, no.
3, pp. 173179, 2010.
39. S. Mishra, A. B. Jha, and R. S. Dubey, Arsenite treatment induces oxidative stress, upregulates
antioxidant system and causes phytochelatin synthesis in rice seedlings, Protoplasma, vol. 248, no. 3,
pp. 565577, 2011.
40. T. Liu, J. Van Staden, and W. A. Cress, Salinity induced nuclear and DNA degradation in
meristematic cells of soybean (Glycine max (L.)) roots, Plant Growth Regulation, vol. 30, no. 1, pp.
4954, 2000.
41. T. Karuppanapandian, J-Cheol Moon, C. Kim, K. Manoharan, W. Kim, Reactive oxygen species in
plants: their generation, signal transduction, and scavenging mechanisms, Australian Journal of Crop
Science, vol. 5, no. 6, pp. 709-725, 2011.
42. Tan, W, Meng, Q. W, Brestic, M, Olsovska, K, & Yang, X., Photosynthesis is improved by exogenous
calcium in heat-stressed tobacco plants, Journal of Plant Physiology, vol.168, pp. 2063-2071, 2011.
35