ROLE OF ATP STATUS IN HEAT STRESS EFFECTS ON PHOTOSYNTHESIS

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: COMPLETE
• Funding Source: NRI COMPETITIVE GRANT
• Reporting Frequency: Annual
• Accession No.: 0192368
• Grant No.: 2002-35100-12057
• Proposal No.: 2002-00732
• Project Start Date: Jul 1, 2002
• Project End Date: Dec 31, 2004
• Grant Year: 2002
• Program Code: [22.1]- (N/A)

Recipient Organization
UNIV OF WISCONSIN
21 N PARK ST STE 6401
MADISON,WI 53715-1218

Performing Department
BOTANY

Non Technical Summary
Reductions of photosynthesis at high temperature are particularly important in limiting or reducing crop yields. Both deactivation of the carbon dioxide fixing enzyme (rubisco) and a reduced ability of the plant to convert light energy into chemical energy by making ATP have been found at high temperature. Rubisco deactivation will cause Rubisco to consume increasing amounts of ATP as the temperature rises. Leaf temperature in crops experiencing heat stress will be measured. We will assess the temperatures experienced by leaves and the frequency and duration of heat stress episodes. The type of heat stress found in field measurements will then be used to design relevant laboratory studies. The relationship between the two effects of heat stress on photosynthesis will be studied by measuring the ATP in the leaf. The plant species used will be Pima cotton (Gossypium barbadense).

Animal Health Component

10%

Research Effort Categories

Basic

90%

Applied

10%

Developmental

(N/A)

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
203	1711	1000	40%
203	1711	1020	60%

Knowledge Area
203 - Plant Biological Efficiency and Abiotic Stresses Affecting Plants;

Subject Of Investigation
1711 - Long fiber cotton;

Field Of Science
1000 - Biochemistry and biophysics; 1020 - Physiology;

Keywords

heat stress

leaf temperature

micrometeorology

adenosine triphosphates

crop yields

stress tolerance

plant response

photosynthesis

plant biology

cotton

plant biochemistry

plant physiology

biochemical mechanisms

rubisco

thylakoids

membranes

interactions

comparative analysis

laboratory tests

leaves

environmental effects

traits

plant improvement

measurement

metabolites

electron transport

fluorescence

carbon dioxide

oxygen

Goals / Objectives
This research will address head on the two most often described mechanisms by which moderate heat stress reduces photosynthesis. On the one hand, the CO2-fixing enzyme Rubisco deactivates. The process of reactivation requires energy in the form of ATP. On the other hand, membranes have long thought to be particularly sensitive to high temperature. If thylakoid membranes are damaged by high temperature, the ability of the plant to make ATP will be compromised. Both effects occur, but their relative importance, and even more their possible interaction, needs to be explored to gain a complete understanding of how high temperature reduces photosynthetic capacity. One of the novel aspects of the proposed work is that the heat stress experienced by individual leaves will be assessed. This assessment will be used to design the stress protocol used in laboratory experiments. It is anticipated that this work will determine how ATP levels change during and after high temperature. Comparing chronic high temperature stress with acute high temperature episodes will broaden the scope the measurements and make the results relevant to more of the experiences of the plant. This work is part of a long-term interest in how the environment affects photosynthetic performance and hence plant productivity. This research may identify specific traits that allow photosynthesis to persist at higher temperatures. This could eventually lead to identifying ways of engineering plants for increased heat tolerance.

Project Methods
The temperature stress experienced in natural conditions will be measured by traveling to cotton fields in Arizona and placing thermocouples on leaves of plants growing in normal cropping situations. The temperatures experienced by leaves in the field will be replicated in a laboratory gas exchange apparatus. This will require mechanisms for changing leaf temperature in the lab as quickly as it can change in the field. We will use infrared radiation from a heat source to rapidly change leaf temperature. Leaf samples will be taken at specific times during the heat stress and metabolite levels, especially ATP, will be measured. In related experiments the rate of electron transport will be assessed by monitoring chlorophyll fluorescence. The effect of heat on the capacity for electron transport will be assessed by manipulating the concentration of carbon dioxide and oxygen in the air surrounding the leaf. These measurements will be compared to determine which processes are most sensitive to heat and how acute versus chronic high temperature reduces photosynthesis. Recovery from the stress will also be assessed by measuring photosynthesis for periods of time after relief of the heat stress.

Progress 07/01/02 to 12/31/04

Outputs
It is well established that adenosine triphosphate (ATP) is a critical energy reserve in living organisms. The hypothesis that ATP levels were reduced by heat stress of plants was tested. We found that ATP levels did not fall even though other changes occurred that should have reduced them. Therefore, we investigated the changes in the plant chloroplasts more deeply. We found that membranes become leaky at high temperature and this leads to increased rates of cyclic electron flow. Surprisingly, we found that the stroma of chloroplasts of heat-stressed plants became more oxidized at the same time that photosystem one became more reduced. This led to a greater understanding of the important role cyclic photophosphorylation has during and after heat stress. From the data, we hypothesize that the movement of electrons from the stroma to the electron transport chain (plastoquinone) is substantially speeded up as a result of heat stress. Understanding the path of these electrons may allow us to predict how to make photosynthesis more robust during heat stress.

Impacts
This information will lead to an understanding of how heat damages photosynthesis and may help find methods for making crops less susceptible to heat damage.

Publications

• Wise RR, Olson AJ, Schrader SM, Sharkey TD (2004) Electron transport is the functional limitation of photosynthesis in field-grown Pima cotton plants at high temperature. Plant Cell Environ 27: 717-724.
• Schrader SM, Wise RR, Wacholtz WF, Ort DR, Sharkey TD (2004) Thylakoid membrane responses to moderately high leaf temperature in Pima cotton. Plant Cell Environ 27: 725-735.
• Sharkey TD (2005) Effects of moderate heat stress on photosynthesis: importance of thylakoid reactions, rubisco deactivation, reactive oxygen species, and thermotolerance provided by isoprene. Plant Cell Environ. In press.

Progress 01/01/03 to 12/31/03

Outputs
Extensive measurements made on cotton growing in agronomic conditions showed that photosynthetic electron transport is the factor that limits cotton photosynthesis at high temperature when plants are grown under normal agronomic conditions. This is in contrast to recent data that appeared to indicate heat damage to a stroma enzyme known as rubisco activase. We showed that there are significant changes in the redox potential of the stroma when cotton leaves are heated. This is correlated with a shift in energy distribution from photosystem 2 to photosystem 1, causing photosystem 1 to speed up significantly in high temperature. It also could cause the reduction in rubisco activase activity that had been reported. These results are important because they reveal what parts of plants need to be altered to make plants more tolerant of high temperature. In the case of heat stress, thylakoid reactions are most sensitive to the heat damage and the changes that occur as a result of the heat damage cause secondary effects, such as the reported effect on rubisco activase. Therefore, efforts to (1) understand the initial damage caused by heat stress on photosynthesis or (2) to look for methods to make plants more heat tolerant need to focus on thylakoid reactions and specifically the speed-up of photosystem 1 at the expense of photosystem 2.

Impacts
This information will lead to an understanding of how heat damages photosynthesis and may help find methods for making crops less susceptible to heat damage.

Publications

• No publications reported this period

Progress 01/01/02 to 12/31/02

Outputs
In the year 2002 we demonstrated that cotton plants experience many short high temperature episodes by measuring leaf temperature in Arizona. Because of this, lab work was focused on how short heating episodes affect photosynthesis. At 42degreesC rubisco was deactivated as reported by other investigators, but after returning to 30degreesC, photosynthesis was limited by the capacity for electron transport, not rubisco activation. We found that the thylakoid membranes, necessary for ATP synthesis, became leaky. Finally, we showed that the damage was worse when leaves were heated in darkness than when they were heated in the light, proving among other things that the damage was not related to photoinhibition. We were suprised that ATP levels did not fall as we had predicted, which has helped us refine the underlying hypothesis; but the essential part of the underlying hypothesis remains intact. This year also was used to develop protocols for measuring electron transport in thylakoid vesicles at different temperatures. Calibration of the oxygen electrode at different temperatures requires a specialized calibration procedure which has been worked out. One paper has been prepared for publication and will be reported next year.

Impacts
This research should lead to a fuller understanding of how heat reduces the capacity for photosynthesis. In particular, we are establishing what modifications will allow plants to photosynthesize better at high temperatures so that, ultimately, yields can be improved in plants that suffer high temperatures.

Publications

• No publications reported this period