other studies build pharmacological evidence for the existence of NOS like enzyme
in various plant species, the purification of relevant protein is still underway [14].
Nonetheless, the identification of two genes in green algae Ostreococcus tauri
and Ostreococcus lucimarinus share approximately 40% homology to animal NOS
genes and also exhibit NOS-like activity [17]. NR-mediated NO synthesis is very
common and known to involve in several physiological processes and plant defense
against biotic and abiotic stress [16]. In several cases NO production in plant tissues
occurs either through nonenzymatic light-mediated conversion of carotenoids or
enzymatic catalysis through NADPH nitrate reductase. NR, in addition to its primary
nitrate (NO_3) oxidoreductase activity, is capable of reducing NO 2 to NO with low
efficacy [18]. Additionally, NO can also be produced through reductive pathways
by assimilatory nitrate reductase, or through the mitochondrial electron transport
system, or from xanthine dehydrogenase/oxidase [15]. The reductive NO synthesis
from NO 2 can occur in cytoplasm, mitochondria, chloroplast, peroxisomes, and the
apoplast of the plant cells [18]. Plant mitochondrial enzymes present in the matrix or
the intermembrane space are also assumed to oxidize L-arginine to NO.
Undoubtedly, NO has the ability to modify the activity of enzymes and some
key signaling components through posttranslational protein modifications including
protein S-nitrosylation, carbonylation, and tyrosine nitration [2, 13]. While the NOmediated protein modifications have been identified for distinct regulatory proteins
such as antioxidant enzymes, there was less information on general mechanism by
which NO is being sensed across multiple plant processes [2]. However, a study in A.
thaliana suggested a unifying N-end rule pathway proteolysis mechanism involved
in NO sensing in plants [2]. Sufficient data placed cGMP, cADPR, L-phenylalanine
ammonia-lyase (PAL), and PR-1 as effectors of NO levels in plants [13].
In the past decade, researchers deployed different methods to elucidate NOdependent processes including NOS/NR activity assays, NO-binding fluorescent
dyes, and various pharmacologic approaches using NO donors including sodium
nitroprusside (SNP) and S-nitroso-N-acetylpenicillamine and NO scavengers 2-(4Carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO), and quantification of effectors by radioimmunoassay or liquid chromatography tandem-mass
spectrometry [9, 15, 17, 19]. The recent use of targeted nanodonors and nanoshutters
has enhanced the specificity while reducing the pleiotropic responses and artifacts
encountered with pharmacological compounds [12, 16]. Numerous genetic studies
have used Arabidopsis mutants with altered nitric oxide levels such as noa1,
nia1nia2, and nox1 to confirm the importance of NO accumulation in response to
various stimuli [17]. Furthermore, the nNOS transgenic lines [20] also underlined the
generation of NO as a vital response for increasing plant adaptation to stress [9].
1.2
Introduction to Secondary Metabolites
Plant secondary metabolites are the compounds that have vital role in plant
interactions and defense. While our understanding on their role in fundamental
plant growth and development is improving, diverse arrays of secondary
598
P. Santisree et al.
in various plant species, the purification of relevant protein is still underway [14].
Nonetheless, the identification of two genes in green algae Ostreococcus tauri
and Ostreococcus lucimarinus share approximately 40% homology to animal NOS
genes and also exhibit NOS-like activity [17]. NR-mediated NO synthesis is very
common and known to involve in several physiological processes and plant defense
against biotic and abiotic stress [16]. In several cases NO production in plant tissues
occurs either through nonenzymatic light-mediated conversion of carotenoids or
enzymatic catalysis through NADPH nitrate reductase. NR, in addition to its primary
nitrate (NO_3) oxidoreductase activity, is capable of reducing NO 2 to NO with low
efficacy [18]. Additionally, NO can also be produced through reductive pathways
by assimilatory nitrate reductase, or through the mitochondrial electron transport
system, or from xanthine dehydrogenase/oxidase [15]. The reductive NO synthesis
from NO 2 can occur in cytoplasm, mitochondria, chloroplast, peroxisomes, and the
apoplast of the plant cells [18]. Plant mitochondrial enzymes present in the matrix or
the intermembrane space are also assumed to oxidize L-arginine to NO.
Undoubtedly, NO has the ability to modify the activity of enzymes and some
key signaling components through posttranslational protein modifications including
protein S-nitrosylation, carbonylation, and tyrosine nitration [2, 13]. While the NOmediated protein modifications have been identified for distinct regulatory proteins
such as antioxidant enzymes, there was less information on general mechanism by
which NO is being sensed across multiple plant processes [2]. However, a study in A.
thaliana suggested a unifying N-end rule pathway proteolysis mechanism involved
in NO sensing in plants [2]. Sufficient data placed cGMP, cADPR, L-phenylalanine
ammonia-lyase (PAL), and PR-1 as effectors of NO levels in plants [13].
In the past decade, researchers deployed different methods to elucidate NOdependent processes including NOS/NR activity assays, NO-binding fluorescent
dyes, and various pharmacologic approaches using NO donors including sodium
nitroprusside (SNP) and S-nitroso-N-acetylpenicillamine and NO scavengers 2-(4Carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO), and quantification of effectors by radioimmunoassay or liquid chromatography tandem-mass
spectrometry [9, 15, 17, 19]. The recent use of targeted nanodonors and nanoshutters
has enhanced the specificity while reducing the pleiotropic responses and artifacts
encountered with pharmacological compounds [12, 16]. Numerous genetic studies
have used Arabidopsis mutants with altered nitric oxide levels such as noa1,
nia1nia2, and nox1 to confirm the importance of NO accumulation in response to
various stimuli [17]. Furthermore, the nNOS transgenic lines [20] also underlined the
generation of NO as a vital response for increasing plant adaptation to stress [9].
1.2
Introduction to Secondary Metabolites
Plant secondary metabolites are the compounds that have vital role in plant
interactions and defense. While our understanding on their role in fundamental
plant growth and development is improving, diverse arrays of secondary
598
P. Santisree et al.