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Nitric Oxide and Programmed Cell Death in Aleurone |
Nitric oxide (NO) is an
important second messenger in animal cells, and accumulating evidence
suggests it is important in plant cells, as well. Exogenous NO has been
shown to affect the responses of plants to pathogens, light, gravity,
and oxidative stress. A paradoxical aspect of NO is that in some
systems it hastens programmed cell death (PCD), whereas in
others, it acts as an antioxidant and delays it. For example,
when suspension-cultured soybean (Glycine max)
cells are infected with the bacterial pathogen Pseudomonas
syringae, NO increases in parallel with other reactive oxygen
species (ROS) and promotes the hypersensitive response and PCD. On the
other hand in potato leaves infected by the pathogen Phytophthora infestans, NO acts as an antioxidant and
inhibits PCD. In this issue, Beligni et al. (pp. 1642-1650)
examine the possible role of NO in mediating GA-induced PCD in barley (Hordeum vulgare) aleurone. They report that NO prolongs the
life of barley aleurone cells incubated in GA. The effects of NO can be
mimicked by the antioxidant butylated hydroxy toluene, indicating that
NO may act as an antioxidant in aleurone cells. They also provide
evidence that aleurone cells synthesize NO, and suggest that NO may be
an endogenous modulator of aleurone cell viability.
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Phytochrome B Mutant with Altered Circadian Rhythm |
Because of the circadian rhythm in the opening and closing of
stomata, plants also show a circadian rhythm in sensitivity to the
toxic gas SO2. Salomé et al. (pp.
1674-1685) took advantage of this fact to screen a population of
mutagenized Arabidopsis plants for individuals that exhibited damaged
leaves in response to SO2 exposure at a time when
wild-type plants were resistant. In this issue, they report that in the
mutant out of phase (oop1) that they isolated by
means of the SO2 screen, the
acrophase peak in several rhythms, including leaf
movement, CO2 assimilation, and
LIGHT-HARVESTING CHLOROPHYLL a/b BINDING PROTEIN
(LHCB) transcription, occurs earlier than in wild type (Fig.
1). The defect in circadian phasing
is seen in seedlings entrained by a light-dark cycle, but not
in seedlings entrained by a temperature cycle. The oop1 mutant also displays a strong photoperception defect in red light characteristic of phytochrome B (phyB) mutants.
Evidence is presented that the oop1 mutation is a nonsense
mutation of PHYB that results in a truncated protein of 904 amino acids. Thus, PHYB contributes information about light
conditions that are critical for proper determination of circadian
phase.
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Figure 1.
A comparison of the free running circadian rhythms
in the production of the photosynthetic protein LHCB in
wild-type Arabidopsis (Col), the out-of-phase mutant oop1,
and the phytochrome mutant phyB. Note that the acrophases in
the circadian rhythms of oop1 and phyB-9 occur
almost 2 h earlier than in the wild type.
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Chilling Injury of Rice (Oryza sativa) Anthers |
Exposure of rice plants to a mildly low temperature (12°C
for 4 d) results in male sterility. The molecular events
underlying this sensitivity are poorly understood. In this issue,
Wen et al. (pp. 1880-1891) report on their cloning of
two novel components of a mitogen-activated protein (MAP) kinase kinase pathway (OsMEK1 and OsMAP1) that are induced in
rice at 12°C. An increasing body of evidence suggests that MAP
kinases play important roles in signal transduction in response to
drought, ROS, pathogen defense, wounding, and/or low temperature in
plants. The components of the MAP kinase cascade include MAP kinase
(MAPK), MAP kinase kinase (MAPKK, also known as MEK), and MAP
kinase kinase kinase (MAPKKK, also known as MEKK). A variety of genes
encoding MAPKs, MAPKKs, and MAPKKKs have been cloned from different
plant species. OsMEK1 encodes for a protein with features
characteristic of a MAPKK. Although OsMEK1 transcript
levels were induced in rice anthers by 12°C treatment for 48 h,
no induction of OsMEK1 transcripts was observed in
4°C-treated seedlings. In contrast, rice lip19, which
encodes for a bZIP protein that is believed to be involved in
low-temperature signal transduction, was not induced by 12°C but was
induced by 4°C. These results suggest that at least two signaling
pathways for chilling stress exist in rice. The MAP kinase
pathway that utilizes OsMEK1 and OsMAP1 as components may
be involved in the sensitivity of rice microspores to mildly cold temperatures.
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Alternate Oxidase and PCD |
Mitochondria play a central role in energy and carbon
metabolism in most eukaryotic cells, being the site of both the Krebs cycle and oxidative phosphorylation. In plants, the electron transport chain (ETC) supporting oxidative phosphorylation branches at
ubiquinone. Electrons flow either from ubiquinone through the usual
cytochrome (cyt) pathway or to an alternative oxidase (AOX). Electron
flow from ubiquinone to AOX bypasses two of three sites of energy
conservation supporting oxidative phosphorylation, thereby reducing the
energy efficiency of respiration. Studies of transgenic plant cells
with altered levels of AOX support the hypothesis that this protein, by
preventing the over-reduction of ETC components, dampens the generation
of ROS by mitochondria.
In animal cells and possibly some plant systems, mitochondria also play
an active role in the process of PCD. A prevalent theme in animal PCD
research is that intracellular redox state may play a critical role in
the overall process. In this case, ROS generated by the mitochondrial
ETC itself may be significant. In this issue, three contributions from
Greg Vanlerberghe's laboratory are devoted to examining the possible
role of the AOX in controlling PCD in plants. Vanlerberghe et al.
(pp. 1829-1842) report that Cys down-regulates the cyt pathway in
tobacco (Nicotiana tabacum) cells. They show that this
down-regulation involves changes in protein phosphorylation and
cytosolic protein synthesis, and is accompanied by an increase in AOX
capacity. The induction of AOX under these conditions still enables the
cells to maintain high rates of respiration, indicating that the lesion
triggered by Cys is in the cyt path downstream of ubiquinone.
Consistent with this notion is their finding that transgenic (AS8)
cells that are unable to induce AOX (due to the presence of an
antisense transgene) lose all respiratory capacity upon Cys treatment.
In AS8 cells, this initiates a PCD pathway, as evidenced by the
accumulation of oligonucleosomal fragments of DNA as the culture dies.
In a second contribution, Robson and Vanlerberghe (pp.
1908-1920) establish that tobacco AS8 cells show increased
susceptibility to three different death-inducing compounds
(H2O2,
salicylic acid, and the protein phosphatase
inhibitor cantharidin). Death induced by
H2O2 or salicylic acid
occurs by a mitochondria-dependent pathway characterized by cyt
c release from the mitochondrion, whereas death induced by
cantharidin occurs by a pathway without any obvious mitochondrial
involvement. The results indicate that plants maintain both
mitochondria-dependent and mitochondria-independent pathways of PCD,
and that AOX may serve as an important mitochondrial "survival protein" against such death.
In the third contribution, Ordog et al. (pp.
1858-1865) take advantage of tobacco AS8 cells to examine the
potential role of AOX in the hypersensitive response, a PCD process
that is associated with the defense of plants against invading
pathogens. This course of research was prompted by previous
pharmacological studies indicating that the AOX inhibitor
salicylhydroxamic acid (SHAM) compromises disease resistance to
viruses. The studies discussed in this issue, however, using transgenic
tobacco plants with altered levels of AOX protein suggest that AOX is
not a critical component of the previously characterized SHAM-sensitive
pathway important in viral resistance.
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Arabidopsis Copper Shuttle Protein |
During their investigations of HR-like cell death induced in
tobacco leaves by the bacterial polypeptide harpin, Balandin and
Castresana (pp. 1852-1857) identified a new cDNA
(AtCOX17), the expression of which is activated by high
concentrations of Cu, by bacterial inoculation, SA treatment, and
treatments that generated NO and
H2O2. All of the conditions
inducing AtCOX17 are known to inhibit mitochondrial
respiration and to produce an increase of ROS, suggesting that gene
induction occurs in response to stress situations that interfere with
mitochondrial function. Sequence analysis of a partial cDNA from
tobacco and that of the corresponding Arabidopsis cDNA revealed
significant homology with COX17, a gene from yeast
(Saccharomyces cerevisiae) and vertebrates encoding a Cu
shuttle protein that delivers Cu to the mitochondria for the assembly
of cyt oxidase. Moreover, the Arabidopsis COX17 cDNA
complements a COX17 mutant of yeast restoring the respiratory deficiency associated with that mutation. These two lines of evidence indicate that the plant protein identified here is a
functional equivalent of yeast COX17 and might serve as a
Cu-delivery protein for the plant mitochondria.
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