Feature: Proteomics on the dark side

This feature appeared in the July/August 2011 issue of Australian Life Scientist. To subscribe to the magazine, go here.

The sun sets over a paddock of wheat in the Western Australian wheat belt. Photosynthesis ceases and, as the moon rises, the growing crop switches rapidly from photosynthesis to dark respiration. In doing so, it begins to exhale carbon dioxide instead of oxygen from its leaf stomata.

According to Professor Harvey Millar, before photosynthesis begins again after dawn, such crops can return as much as 70 per cent of the carbon they captured from the atmosphere the previous day. The rest is retained as biomass: cellulose, lignin, starch, oils and sugars.

Millar says understanding what happens during the transition from photosynthesis to dark respiration and back again is crucial to accurate modelling of global carbon storage by plants, particularly as carbon dioxide levels in the atmosphere rise to the highest levels in the past 20 million years.

Tilting the balance between the two competing processes to increase carbon storage by just a few per cent could be doubly beneficial: it could increase crop yields, especially for the ‘big three’ global crops of wheat, rice and maize by millions of tonnes; meanwhile sequestering more carbon dioxide from the atmosphere in the form of biomass.

The biochemical reactions involved in energy production and respiration take place in mitochondria in the plant’s leaves and green tissues.

“We don’t know much about how respiration rate is actually determined, even though we’ve known for a long time that plants markedly change their respiration rate during the day-night cycle,” says Millar.

“Nobody believed the proteome could be changing rapidly enough to be a significant factor in the change in respiration. It was assumed to be a simple matter of substrate supply and demand.”

A PhD student in Millar’s laboratory at the University of Western Australia, Chung Pong Lee, set out to investigate the mitochondrial events that underlie the transition from photosynthesis to dark respiration and the accompanying changes in respiration rate, and made a remarkable discovery.

“It was a demanding work, involving purifying mitochondria from the leaves of growing plants and analysing the proteome at timed intervals throughout the day-night cycle,” says Millar.

“What we saw was a change in protein content and changes in the composition of the proteome. Some proteins were effectively morning proteins, midday proteins and evening proteins, that cycled in response to external cues, changing entire biochemical pathways. It seems mitochondrial function is remodelled every day and night.

“Leaves are very dynamic, constantly changing their proteomes. Those early findings have taken our research into a whole new area of research into protein degradation, and beyond, using the latest proteomics tools to measure rates of protein degradation.”

Millar’s team is exploring how gene transcription is coordinated during the different phases of protein synthesis and how mitochondria organise the degradation of redundant proteins during the transitions from photosynthesis to dark respiration and back to photosynthesis after dawn.

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Active regulation

Early studies left unresolved the question whether the proteins in mitochondria are synthesised de novo or are recycled when redundant proteins are degraded during the day-night cycle.

Millar says recent work has shown that, in fact, both processes operate, and cells actively regulate the balance between synthesis and breakdown.

“If these processes are under active rather than passive control, it may be feasible to manipulate rates of protein synthesis and degradation,” he says.

“You can’t expect to make gross changes to the way plants respire, but it may be possible to subtly modify the rates at which individual proteins are synthesised or broken down, and do it in a tissue-specific way.”

“Reducing the net loss of carbon dioxide through respiration from leaves over the lifetime of plants is probably our main objective,” he says.

“Dark respiration occurs on a staggering scale. Globally, plants respire six to eight times more carbon dioxide at night than all the carbon dioxide emissions from the burning of fossil fuels.

“So a relatively small reduction in the rate at which crop plants respire could have a significant effect on the global carbon cycle. The question is whether we can change it.

“The other issue is what happens to plant respiration as atmospheric CO2 concentrations and global temperatures change. What is the impact on plant structures, which could have a much bigger impact on capacity of plants to take up excess CO2 from fossil fuel burning?”

Millar says researchers modelling the role of plants in the global carbon cycle, and their response to global warming, are taking “a lot of interest” in Australian research on plant respiration.

Respiration rate also turns out to be influential in another key environmental determinant of wheat yields: salinity tolerance. “A wheat cultivar’s salt tolerance is measured by comparing its ability to produce biomass in saline versus non-saline conditions. Salinity tolerance is defined as the ratio between the two.”

Millar says respiration rate may be a significant part of the equation: a variety of plants increase their respiration rate under salty conditions, because excluding salt from the roots is an energy-intensive process. The extra energy demand reduces the amount of photosynthate the plant can allocate to biomass, slowing its growth.

Read part II: Proteomics and plant respiration.