Annual Report -1996 Regional Research
Annual Progress Report:
Project Title:
Characterization and Mechanisms ofPlant Responses to Ozone in the
Coonerating Agencies and Princinal
Investigators/ Renresentatives:
State
or A2encI PI/ Renresentative
USDA -ARS
(Beltsville, MD)
M.
Robinson USDA -ARS & N.C. State U. (
J. Miller
*** USDA- CSREES J. Barnes ** US Environmental Protection Agency (
~* CSREES
Advisor
~**
Research Leader and CRIS Project Lead Scientist at the participating USDA-ARS
Research rpntpr~
PROGRESS AND ACCOMPLISHMENTS IN 1996
Several projects were
conducted in 1996: A study was completed and analyzed on the
detection of ozone effects on the response of
loblolly pine grown in a competitive environment with other woody and
herbaceous species. Results indicated the greatest reduction in pine gro' at sub ambient levels of ozone. Species numbers and
percent cover were the greatest for this treatment and were greatly reduced
with increasing concentrations of ozone. A MS thesis was completed on this work
and publications are currently being written. Exposure-response relationships
between several native plant species sensitive to Ozone and Ambient Ozone
concentrations in Class I Wilderness areas throughout the South are currently
being developed Tall milkweed was inventoried for ozone sensitivity for the 5th
year in the Great Smoky Mountains National Park. Injury increased with time and
varied by location. A study was completed on the sensitivity of several
southern plant species to ozone. Most species tested WI intolerant to ozone.
Butterfly bush, however, was relatively sensitive to ozone.
Maryland -U Diversity of
Maryland, College Park, Agronomy Department in Collaborat
with USDA -ARS, Beltsville, MD, Climate Stress Laboratory (CSL), Remote Sensing
an Modeling Laboratory (RSML) and Electron Microscope Laboratory (EML) -Dr. Charl. Mulchi, Drs. Steven Britz, Craig Daughtry
, Edward Lee, Michael Robinson, and Williall Wergin.
Three field investigations
were performed in 1996 involving open-top chambers, one involving wheat and two
for soybeans. The wheat study consisted of growing two cultivars (Gore and
Susquehanna) at two levels ofC02 (355 vs. 500 nmo1/mol C02) and two 03 regime:
(charcoal filtered vs. nonfiltered air + 35 ,umo1/mol
03) under well-watered conditions. Irregl stands in
several sheltered plots caused the low moisture treatments to be dropped for
1996. 1 photosynthesis, cWorophyll fluorescence and
leaf area index values were determined at
preflowering, anthesis and
grain fill. Also, vegetative samples (i.e. flagleaf)
were collected dun each of the three periods and frozen in liquid N2 for enzyme
assays concerning several antioxid enzyme systems.
Biomass and grain samples were collected at maturity.
Soybeans were grown following the wheat
involving a similar 2 x 2 factorial of C02 anc 03
treatments and two moisture regimes (well watered vs. sheltered plots).
Photosynthetic ratl stomatal characteristics, cWorophyll fluorescence and LA! data
were determined throughout th growth of the plants.
In addition, canopy spectral reflectance characteristics were determined c 250
wave bands over the range 400 to 1100 ,um in
an effort to develop remote sensing procedures for plants under interactive
stress levels. Grain yields and quality characteristics W{
measured following plant maturity.
The
second soybean investigation concerned the screening of 22 cultivars over maturit) groups III, IV, and V for 03 tolerance. Ten plants
of each cultivar were grown in OTC's expo to CF and NF + 03 (ie 70 mol/mol O~ ) for 28 days.
There were four replicates of each treatment. Three plants were harvested at
time zero when treatments wre intitiated
and
were harvested 19 days later. Leaf areas,
leaf and stem weights were measured on the three plants from which relative
growth rates and relative leaf extension rates were calculated. Leaves from the
remaining plants were sampled for antioxidant enzyme assays. The theory behind
this work is that there should be a correlation between changes in growth
activities and antioxidant enzyme activities with treatments. The goal of the
study is to perfect a seedling screening procedure which accurately reflect the
ability of cultivars to detoxify 03. This work will be continued and expanded
in 1997.
Work continued on
assessment of the effects of ozone on growth and development of white clover
clones (Trifolium repens)
NCS (ozone-sensitive) and NCR (ozone resistant), originally obtained from
A.S. Beagle, USDA / ARS, Raleigh, NC, focusing on responses at 7 and 28 day
intervals. Growth of rooted cuttings was compared in natural soil (NS) and
artificial growing medium (AGM) in greenhouse chambers for 28 days at 80 ppb
ozone, 7 hours/day.
ACM plants were fertilized weekly, NS
plants were not fertilized. ozone reduced root and
shoot weights for NCS and NCR in AGM, but only for NCS in NS. Flower numbers
and weights were reduced by ozone for NCS and NCR in AGM, but not in NS. Ozone
reduced total stolon weight NCS in both AGM and NS,
but not for NCR. NCS was affected by ozone in AGM and NS,
while NCR was only affected in AGM. Foliar
biomass response of potted NCS and NCR plants, with and without weekly sprays ofEDU (ethylenediurea) (300 ppm) to ambient ozone was
determined in the field under ambient ozone conditions. Ambient ozone
concentrations were lower than usual, resulting in similar values for EDU and
clone interactions, with detection of an ozone effect in NCS by means ofEDU treatment at the fourth harvest only. NCS and NCR
plant: were also planted directly into the ground in a field plot. Unlike
potted plants, these were not watered of fertilized or sprayed for insects.
Three biomass harvests were obtained. NCS grew faster and produced more biomass
than.NCR all season. EDU had no effects on NCR. EDU
increased growth ofNCS at harvests 2 and 3. These
results suggest that comparisons should be made between response of potted and
planted clover in the field.
Previous work suggested
that mid-range ambient ozone concentrations (50- ~70 ppb) ma) be important in
causing crop yield loss. To further examine this observation, the relationships
between ambient 03 concentrations and key meteorological variables that promote
03 flux and plant uptake were examined at 6 geographic locations varying in
their elevations above the sea level. These locations were: Beltsville, MD (39
m MSL), Bondville, IL (212 m MSL), Georgia Station,
GA (270 m MSL), Perkinstown, WI (472 m MSL), Big
Meadows, V A (1073 m MSL), :tnd Grand Canyon, AZ
(2085 m MSL). The daylight (global radiation ~ 50 W m-2) hourly
:tmbient 03 concentrations for 1989 -1994
growing seasons were stratified by their range and their frequency
distributions were related to varying ranges of air temperature, water vapor
pressure deficit and mean horizontal wind
velocity. High 03 concentrations (>70 ppb) normally coincided with high air
temperatures (>20° C) and therefore high water vapor pressure deficit
(>20 hPa). Similarly, high 03 concentrations often
coincided with low to moderate horizontal wind velocities (1 -2 m sec -1). Such
high concentrations also co-occurred with moderate to lov
atmospheric conductivities ( a product ofmean horizontal wind velocity, global radiation and ail
temperature) .The diurnal patterns of atmospheric conductivity and other 03
uptake limiting factors (e.g. vapor pressure deficit) lead to a much greater
probability for a high 03 uptake from early to mid-morning to mid-afternoon
hours than in the later part of the day.
Our research efforts
continued on transgenic Bel W3 tobacco plants which
were generat{ in our laboratory to express altered levels of antioxidant
enzymes. Bel W3 lines which were derived from several
independent transformants constructed to over express
ascorbate peroxidas (APX) in the cytosol were tested
for tolerance to acute ozone exposures. These lines never showed protection
from ozone injury, while in contrast they were very tolerant of oxidative stres generated by the redox-cycling herbicide, paraquat. Rather, several transgentic
lines showed more injury than their respective nontransgenic
controls and this was particularly striking in your leaves. In addition, in
collaboration with Dr. Pell's laboratory, we showed that overproduction ( APX in the chloroplast did not protect Bel
W3 tobacco from ozone injury. Lastly, transgenic B{ W3
tobacco plants which over express or under express monodehydroascorbate
reductase (MDAR) were generated and examined for sensitivity to ozone. The
progeny that overproduce or sense-suppress MDAR responded similarly to nontransformed controls to exposures of acute ozone.
Sugar Maple Research
Experimental fumigations
of sugar maple trees were conducted in 1995 using both open- top chambers and
an open-field exposure system. Two separate experiments were conducted. 1 the
first, led by Mary Topa, three year old seedlings
from 5 different families were exposed in open-top chambers to ambient ozone
and 1.7 and 3 times ambient ozone under two light regime~ (15 and 35 percent offull sun). The experiment is a split-plot with ozone as
the main plot and shading as the sub-plot. The objective of the research id to
measure ozone-induce changes in carbon allocation. This was the final year of
exposure and the data are not yet summarized.
In the second experiment,
led by David MacLean, Jay Jacobson, and Jonathan Comstock sugar maple saplings
(16 years old and approximately 10 meters tall) were exposed to ambient 0] 3X
ambient ozone for the fourth growing season using an open-field fumigation
system constructed in and around a small stand of trees. Trees were not exposed
to elevated ozone
During other times saplings were exposed
to ambient ozone. The treatments had a pronounced ~ffect
of leaf carbohydrate levels and on carbohydrate partitioning between soluble
sugars and ;tarch. Very
similar patterns were seen each year between 1994 and 1996. In general, total
1onstructural carbohydrate (TNC) tended to be lower in the 3X ambient ozone
treatments due to iepression of starch pools. Early
in the season, sugar pools were actually elevated under 3X imbient ozone, compensating for depressed starch and
resulting in similar TNC levels. As the ;eason
progressed, sugar pools became similar between high and low ozone treatments,
while ;tarch pools and total TNC became more and more
depressed in 3X ambient ozone treatments. Starch: Sugar ratios were lower under
3X ambient ozone on all sampling dates. Diurnal [luctuations
in TNC pool sizes also ceased earlier in the season in 3X ambient ozone
treatments. rhe differences in carbohydrate pool
sizes and partitioning were not associated with consistent iifferences
in leaf gas-exchange rates as measured with a LICOR 6200. A comparison with
Jotted seedlings exposed to the same
treatments in open-top chambers in 1994 and 1995 showed i similar trend, but
the effect was much less pronounced in seedlings.
Research to identify
native bioindicator species for ozone and use them in field surveys has )een conducted at
hat species, and the environment must allow
gas exchange to occur so ozone can be taken up by he plant. The interaction of
the last two criteria is particularly important since it determines the ~ffective dose of ozone the plant will receive. The studies
at
Whole plant responses to
ambient ozone on
Research Activities and
Findings: We have been investigating the chronic effects of elevated ozone and
carbon dioxide on the growth and physiology of eastern white pine and
yellow-poplar in open-top chambers for the last five years.
Three major hypotheses
were tested: 1) C02 enrichment ameliorates the negative effect~ of 03; 2) the
relative response to 03 and C02 is the same in short-term studies with
seedlings as with older trees; and 3) faster growing hardwood species are more
sensitive to 03 and more responsive to elevated C02 than slower growing conifer
species.
Study design: Within each
plot, 12 plants were arranged in a circular pattern approximately 1.8 m in
diameter. The white pine plantation consisted of seedlings from two sources: 1)
northern genotype (2-0 stock) from the Upper Peninsula ofMichigan
(Baraga County), obtained from Dr. David Karnosky, Michigan Technological
University, Houghton; al 2) southern genotype (1-0 stock) from northern Ohio
(near Toledo), obtained from the State Nursery of Ohio, Marietta. Six seedlings
of each source were planted within each plot. The yellow-poplar seedlings were
1-0 stock obtained from a private nursery in western Pennsylvani.
Seed collections were made in southeastern
charcoal filtered air (CF); ambient 03
concentration (IX); two times ambient 03 concentration (2X); two times ambient
03 concentration plus 350 ppm C02 above ambient (2X + COJ; and open-air,
chamberless plot (OA). The 03 and C02 were dispensed automatically 24 hours per
da from mid-May through mid-October in 1992 through
1996. Standard 3m diameter open-top chambers were used, In
1994, open-chambers (yellow poplar only) were modified to increase their height
to 4.6m. In 1995, these chambers were replaced with 4.7m wide and 10m tall
large tree open-top chambers to accommodate the increasing size of the
saplings. Monthly growth aru physiological
measurements taken during each growing season included stem height and basal
diameter, photosynthesis, stomatal conductance, chlorophyll content, and foliar
nitrogen and phosphorous concentration. Additional monthly foliar collections ofboth species were made in 1995 and 1996 to monitor
treatment impacts on total non-structural carbohydrates. However, due to
limited funding, these samples have yet to be analyzed. If a proposal submitted
with Drs. Weinstein and RetzlaffofBTI to NlGEC is funded, the carbohydrate analysis will be done in
1997. In late September 1993, shoot systems of6 yellow-popular seedlings from
each chamber were destructively harvested and leaf area and leaf, stem and root
dry mass determined. Two additional destructive harvests (subsamples
of three saplings per plot per harvest date) were mac
in September 1994 and September 1996. White pine were
destructively harvested in 1994 and 1996 (6 seedlings/chamber/harvest). We are
currently excavating root systems ofboth species.
Prior to 1995, very little
direct impact of ozone on the growth and biomass production o yellow-poplar was
observed, although photosynthetic rates were decreased by ozone. Significan increases in height and diameter growth ofyellow-poplar-exposed to ozone plus elevated carbon
dioxide were observed throughout the study. In october 1995, for the first time in this study,
reductions in total height and diameter growth were observed for saplings
exposed to ozone. Total stem height was 633, 583, 586, and 682 cm for
yellow-popular grown in CF, IX, 2X, and 2X + C02-air ,
respectively (p = 0.01). Stem diameter was similarly affected by ozone and
elevated carbon dioxide (p = 0.03). Ozone-induced growth reductions appear to
have been ameliorated by the elevated carbon dioxide in the 2X + C02 treatment.
Throughout the study, photosynthetic rates were stimulated by exposure to
elevated C02 + 03. By late 1994, lowered photosynthetic rates of trees grown in
2X + C02-air suggest down-regulation or acclimation to elevated carbon dioxide
may have been initiated. This same trend was not consistently observed during
the 1995 season, but preliminary analysis of the 1996 photosynthetic data
indicate that photosynthetic acclimation may be occurring.
Throughout most of this
study, white pine has not been very responsive to the experimental treatments.
For the most part, no seed source treatment effects were observed. Preliminary
analysis of the white pine growth data in 1996, show no significant 03, or 03 +
C02 effects.
Experiments were conducted
to test the hypothesis that °3-induced accelerated foliar senescence in hybrid
poplar provides recyclable nitrogen to the plant when soil is nitrogen limite To test this hypothesis
hybrid poplar cuttings were grown in sand culture with a modified Hoagland's
solution. Nitrogen levels were initially set at 50 ppm. At six time points
during the experiment ramets were harvested for total
biomass and growth analysis. Prior to the harvest foliage were subjected to gas
exchange analysis and then harvested to assess specific measures c senescence
and nitrogen partitioning. The first harvest was conducted at Julian date 192.
At Julian date 203 a subset of plants had nitrogen addition reduced from 50 ppm
to either 25 ppm ( 10 ppm; an additional subset of plants were subject to a
withdrawal from the 50 ppm nitrogen supplementation to 25 or 10 ppm at Julian
date 215. Data analysis is in progress. It appears thc
while 03 initiates accelerated senescence in all treatments, the percent of
leaves abscised is high( in plants from which 80% of
the nitrogen was withdrawn on Julian date 203.
Experiments were conducted
to test the hypothesis that plants grown in high light would be more tolerant
of 03 because they would have higher levels of antioxidant activity. Wild grap' plants were grown in open-top chambers either in full
sun or in continuous shade. Plants were harvested periodically for growth
analysis. In addition to gas exchange and chlorophyll fluorescence analyses
were conducted nondestructively periodically throughout the experiment. Foliage
were harvested for analysis of antioxidant enzymes.
Data analysis is incomplete at this time but preliminary results do not suggest
an O3-light interaction.
Studies have been
initiated to develop a greater understanding of the mechanism by whi( 03
induces stomatal closure. Experiments are planned to conduct ecophysiological
measurements, by conventional patch clamping and laser patch clamping of guard
cell protoplas and interact cells, respectively, to
characterize the effect of 03 on ion pumps and channels. In preparation for
these studies we have treated isolated epidermal peels of Vicia
faba with 03, either
applied in the gaseous phase, or in solution, and observed guard cell
performance. Ozon( induces stomatal closure just as occurs in situ. This
suggests that 03 -induced stomatal closure can occur in the absence of
mesophyll effects.
Studies have been
initiated with Arabidopsis. An °3-induced ACC synthase cDNA has been cloned from Arabidopsis foliage.
This cDNA is different from the two ACC synthase
cDNAs previously cloned from 03-stressed potato foliage. Transgenic Arabidopsis
plants carrying promoters for two senescence associated genes (SAG 12 and SAG
13) which have bee fused to GUS constructs (provided by R. Amasino
), were treated with chronic doses of 03, at levels which did not elicit
visible injury. The plants with the SAG 13 promoter, which codes for an alcohol
dehydrogenase promoter, exhibited GUS activity in foliage after a brief
exposure to ( and well before it was expressed in nontreated tissue. The plants with the SAG 12 promoter,
which has high homology for a cysteine protease
promoter, also exhibited GUS activity more rapidly in 03-treated plants, but
not as early as the SAG 13 plants. Much more analysis of this plant material and
these experiments is underway. These data provide evidence that genetic
regulation of 03-induced accelerated senescence occurs in the absence of foliar
injury suggestin!
A field experiment
initiated in 1995 using open-top chambers was repeated for a second season in
1996 in order to evaluate the role of flavonoids in sensitivity to ozone.
Soybean (Glycine max [L.] Merr.) isolines either lacking flavonoids
or containing different combinations kaempferol
glycosides were compared. Previous experiments using greenhouse-grown plants
exposed briefly to high levels of ozone showed that one line (OX941 ) was significantly less sensitive to ozone than
related lines. OX941 contains a flavonoid (K9) that is associated with reduced
numbers of stomata, reduced photosynthetic rates, lower yields, and increased resistanc to drought. In 1995, physiological (stomatal
conductance and photosynthesis) and yield measurements in the field confirmed
that OX941 was behaving as expected. Moreover, chronic ozone exposure had
little influence on the already low yield of this isoline.
However, leaf chlorophyll measurements and subjective observations of damage
also indicated that OX941 wa
more severely damaged by ozone early in the season than any of the other lines.
These observations suggested that K9 may have effects on ozone sensitivity
unrelated to any effects or stomatal conductance. Data from 1996 are largely
consistent with previous results.
A study was undertaken to
compare the influence of ozone on the total vitamin C ( ascorbate +
dehydroascorbate) levels and ascorbate to dehydroascorbate redox ratios in
mature leaflets of tv soybean cultivars were grown in
the field concurrently in carbon-filtered, open-top chambers (3: ppb ozone) and
non-filtered open-top chambers supplemented with additional ozone (60 ppb
ozone). Since glucose is the precursor of ascorbate, the daily rate of net
disappearance of glucol and the daily rate of net
accumulation of glucose sources including sucrose and starch, was monitored
concurrently. It was concluded that cv Essex was more ozone-tolerant than cv Forre: in part because leaflets of cv Essex were able to
maintain a higher ascorbate to dehydroascorbat ratio
and a higher total vitamin C (ascorbate plus dehyroascorbate)
level during exposure ofplal to elevated ozone.
However, in leaflets of both cv Essex and cv Forrest plants, levels of glucose
and glucose sources such as sucrose and starch appeared to be adequate to
support ongoing ascorbate synthesis, even during long-term chronic exposures to
elevated ozone.
A cooperative field
experiment between theCSL and the University ofMaryland,
North Carolina -USDA-ARS & N.C. State
University -Drs. F.L. Booker, A.S. Beagle, J.] Miller, K.O. Burkey, E.L.
Fiscus, C.D. Reid, RA. Reinert, and S.R. Sharer
Factorial dose-response
studies were performed in open-top field chambers to determin~
the agricultural importance of interactions between current and expected
concentrations of 03 a CO2. Cotton, soybean, and rice were exposed from shortly
after emergence to maturity, to a range ofC02 concentrations (1.0 to 2.0 times
ambient), combined with a range of03 concentrations (0.5 to 1.5 times ambient).
Biochemical and physiological responses were measured periodically and yield
was measured at maturity .C02 enrichment caused positive growth responses of
the three crops but the apparent C02 effects were greater when plants werc stressed by 03 than when not stressed by 03. For
example, at 0.5 times ambient 03 (non-stresse
plants), C02 enrichment did not increase seed cotton yield; at ambient or 1.5
times ambient 03 (moderate to severe plant stress), C02 enrichment markedly
increased seed cotton yield. Result show that the level of 03 stress must be considered to properly interpret plant
response to C02 enrichment. Knowledge of such interactions will allow estimates
of the effects of C02 enrichme for plants at all
levels of 03 stress.
Successful production of
stress-resistant crops and prediction of future agroecosystem productivity
requires identification of key physiological processes that are affected by
stresses induced by climatic changes. Analysis of the limitations to
photosynthesis in soybean using NC curves and biochemical analysis
showed that in elevated C02 (with or without 03), RuBP regeneration was the
major limitation during the vegetative stage and that this limitation disappeared
in the reproductive phase. Elevated C02 alone had no effect on RuBP pool size bu did ameliorate the 03 -induced decrease in the pool.
Experiments with three rice lines showed a average
increase in vegetative biomass of 18% in charcoal filtered (CF) air at double
ambient C02, while a 1.5X ambient 03 treatment resulted in a 24% suppression at
ambient C02. In a C( X 03 treatment, the biomass
suppression by 03 was completely eliminated by double ambient C02. Six snap
bean genotypes, with varying sensitivity to 03, were examined to see if the
sensitivity might be related to differences in 03 uptake.
Generally, 03 caused a decline in leaf conductance that was proportional to
injury, suggesting that changes in leaf conductance were t result of injury and
not a factor controlling the level of injury .An observed doubling of total
phenolics and proanthocyanidins under elevated C02, 03 and low N in cotton may
affect nutrien cycling and plant-pest interactions.
These results indicate that changes in the gaseous atmosphe
environment must be considered in future crop production scenarios associated
with global change.
Studies conducted during
1996 continued to focus on the linkage between carbon allocation, root
metabolism, and soil food-web organisms. In two separate studies we have foun that ozone exposure increases below-ground 02
consumption, CO2 production, and RQ (C02/O2 ratio ).
In both studies, nutrient stress magnified the response to ozone stress. The
question remains as to whether the increase is due entirely to
altered root metabolism, or due to a change in activity offood-web
organisms. We have found that populations ofbacteria
and fungi tend to increase in soils of ozone-treated plants, however
variability in response is high throughout the season.
In growth chamber studies
using wheat as a model system, we have found that short-tern exposure to ozone
increases root exudation, which could stimulate rhizosphere activity and
account for increased soil gas fluxes that have been observed. Whether or not
long-term exposures will result in similar patterns of response remains to be
determined.
Ozone exposure to mixtures
of ponderosa pine and wild blue rye grass in open-top chambers also has
resulted in increased C02 flux from soil. In these systems, we find greater soi] organic matter contents in our ozone treatments than
in controls. Associated with increased soil organic matter, we have measured
increased bacterial and fungal population, although the response is not
consistent throughout the growing season.
Based on our results and
those in literature, we are examining the following as a plausibl{ sequence of events
for ozone exposed plant-soil systems: Decreased carbohydrate allocation below
ground in seedlings exposed to ozone results in less carbohydrate availability
for growth and maintenance of roots. Root exudation increases due to membrane
leakiness and subsequent breakdown, which stimulates rhizosphere activity. RootTNC also is reduced in ozone exposed plants, increasing
rates of root mortality and turnover. We hypothesize that in ozone-exposed
systems, soil food-web organisms are initially stimulated as a result of
increased substrate availability, however, over time,
food-web structure deteriorates as root biomass decreases. Our future studies
are designed to quantify rates of root turnover and soil food-web structure in ozon stressed systems.
Virginia- Virginia
Polytechnic Institute and
Soluble protein
populations and antioxidant enzymes in two clones of white clover (Trifolium repens), cv. Regal,
were examined in response to 03 to determine biochemical mechanisms involved in
oxidative tolerance. The two clones (NC-R, 03-resistant and NC-S, 03-sensitive)
were obtained from A.S. Heagle at USDNARS,
28 day
growing period.
Tolerant and sensitive clones were exposed to a step-function 03 profile (0.50
– 150 ppb maximum concentration for 3h) for 3 days, 8 hr/day in CSTR chambers.
Superoxide dismutase activity of the
three isoforms found in white clover were not
different in NC-R and NC-S prior to 03 exposure, nor did activity of any
isozyme change after ~ days of fumigation. Catalase activity increased in both
clones, compared to control, non-fumigated plants, after 3 days. This increase
was somewhat greater (24%) in NC-R than in NC-S (14%).
Two proteins, detected by two dimensional
gel electrophoresis, began to appear in the tolerant clone after one day of 03 exposure and increased in concentration throughout the
fumigation period. In NC-S, these proteins accumulated to low concentrations,
compared to NC-R, after 3 days of 03 treatment. The
estimated molecular weights of the polypeptides are 2, KD and 27.5 KD and the isoelectric points are 4.1- 4.3. Anion exchange
chromatography oflea extracts from 03-treated clover
revealed the presence of a unique peaks in the
tolerant clone onJ Efforts are continuing to purify
sufficient protein for amino acid analysis.
Importance of 1996 Research
During 1996, members of
the NE-176 committee continued to contribute to a data base which is making it
dramatically clear that in
Plants are often subjected
to multiple environmental stresses, e.g. heat, cold, drought, etc In 1996,
NE-176 members continued studies to examine the interaction of ozone-induced
stress with the effects of other stress components, e.g. drought stress and
competition. This research also will help in the prediction of plant response
during prolonged periods of ozone exposure.
Additionally, NE-176
researchers are finding that conferral of tolerance of crop plants to
ozone-induced oxidative stress is associated with activities of various
antioxidant enzymes, e.g. ascorbate peroxidase and superoxide dismutase. This
knowledge will lead to the understanding ( how to
genetically engineer crop plants which are more strongly tolerant of
atmospheric pollutants. Further, research by NE-176 members has indicated that
elevated atmospheric CO2 ameliorated ozone-induced plant stress. Mechanisms by
which this occurs are being investigated Some recent findings suggest that
growth of plant in elevated atmospheric CO2 results in increasc
antioxidant enzyme activities which are, in part, responsible for conferring to
crop plants additional tolerance to ozone-induced stress.
Finally, NE-176
researchers ha,ve found
evidence that exposure of plants to ozone accelerates leaf senescence, and this
appears to be caused by increased ethylene production in tht
leaves. Prevention of ozone-induced leaf senescence may be facilitated by regulation
of the expression of genes that control the synthesis of enzymes associated
with ethylene biosynthesis. This may represent an inroad into genetically
engineering plants to be able to withstand ozone- induced oxidative stress.
Work Planned for 1997
The proposal for
continuation of the regional project entitled: "Characterization and
Mechanisms ofPlant Responses to Ozone in the
Northeast" has been revised very thorougWy, a
has been approved for a duration of 5 years (1997
through 2000). The research over the next fi years
will use and continue to build upon the research completed during the previous
5 years (1991 through 1995). The program executed during the next five years
will continue to emphasize research across the continuum from molecular/physiological
mechanisms to field responses.
APPROVED:
k~lka"1 a;r"';--L' . ((.. ~ 7'//~/9Z
A.H.
Chappelka Date 1996 Chair, Technical Committee
Patrick.
A
PUBLICATIONS
Journal Articles
Andersen,
C.P., and C.F. Scagel, 1996.
Nutrient availability alters below-ground respiration of
ozone exposed ponderosa pine. Tree Physiol. : (In press).
Andersen, C.P ., R. Wessling, M. Plocher,
and W .E. Hogsett, 1997. Carry-over effects of ozone on ponderosa pine root
growth and carbohydrate concentrations. Tree Physiol. :(In press).
Anttonen, S., M.-L. Sutinen, and A. S. Heagle. 1996. Ultrastructure and some
plasma membrar characteristics ofozone-exposed
loblolly pine needles. PhysiologiaPlantarum
98:309-319.
Booker,
F. L., S. Anttonen, and A. S. Heagle. 1996. Catechin, proanthocyanidin and lignin contents of
loblolly pine (Pinus taeda L. ) needles after
chronic exposure to ozone. New Phytologist 132:483-492.
Brown, P.S., D.P. Knievel and E.J. Pell.. 1996. Drought
effects on ascorbate peroxidase and glutathione reductase activities in
mesophyll and bundle sheath cells of maize. Physiologia Plantarum 95:274-280.
Eason,
G., R.A. Reinert, and J. E. Simon.
1996. SO2-enhanced phytotoxicity of 03 to watermelon. Journal
of the American Society for H orticultural Science.
12 ( 4) : 718- 721.
Eckardt,
N.A. and E.J. Pell.
1996. Effects ofEDU on ozone-induced acceleration offoliar senescence in potato (Solanum tuberosum L.)
Environmental Pollution 92:299-306.
Findley,
D.A., G.J. Keever, A.H. Chappelka, C.H. Gilliam, and D.J. Eakes. 1996. Sensitivityof
six red maple cultivars to acute ozon~ exposures. J Arboricul. 22:264-269.
Findley,
D.A., G.J. Keever, A.H. Chappelka, C.H. Gilliam, and D.J. Eakes. 1997. Ozone sensitivity of selected
southeastern landscape plants. J Environ. Hort. 15:51-55.
Fiscus,
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R., C.L. Mulchi, E.H. Lee, I.N. Forseth and L. Slaughter. 1996. Use od 13C and 1sN isotopes to investigate 03 effects on
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C.P., and P. T. Rygiewicz, 1996.
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presentation, 31 st
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Barbo, D.N. 1996.
Influence of ozone on the productivity and diversity of an early successional
forest community. MS Thesis,
Brendley, B.W. 1996.
Ozone-induced changes in the status of Rubisco in aging foliage of Populus and
Quercus and the role of Rubisco in stress compensation. Ph.D.
thesis. The
Murthy, S.S. 1996. Molecular cloning and analysis ofpea
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