In a eureka moment they discovered that the secret lay in the properties of the mitochondria, the small energy producing organelles in all cells. The respiration rate of the mitochondria from cold-resistant organisms was more sustainable at lower temperatures than the respiration rate of the mitochondria from cold-susceptible organisms. They went on to show that this was due to differences in the lipid composition of the mitochondrial membranes and that similar phenomena occurred in hibernating animals.
The practical applications of this work range from better plant production to preservation of organs for human transplantation.
- How do some living organisms resist the cold better than others?
- Studies with plant mitochondria - a eureka moment
- Extension to animals
- The cause of the rapid decline in mitochondrial respiration rate in cold-sensitive organisms
- Hibernating animals modify their membrane lipids
- Practical applications in plant production and medicine
For many years scientists around the world were puzzled by the ability of certain plants and animals to tolerate low temperatures without distress, while others suffered injury and even death when exposed to the same temperatures. Why do cabbages and broad beans flourish in temperatures far below those which kill melons and tomatoes? How do reptiles, amphibia and fish survive temperatures lethal to many warmblooded animals? Why are some warm-blooded animals able to hibernate, and others not?
The key to the mechanism that enables some plants and animals to resist cold has been found in the myriad cells of which they are composed---more particularly, in the physical properties of the cell membranes. This discovery stems from research on mitochondria – minute particles embedded in the jelly-like cytoplasm that fills each cell. Enzymes associated with the membranes of the mitochondria help break down sugars into carbon dioxide and water and in doing so release considerable amounts of chemical energy – a process known as cell respiration.
In 1968, Dr John Raison of the CSIRO Division of Food Research, who by then was devoting most of his research to mitochondria, was joined in Sydney by Dr James Lyons, Head of the Department of Vegetable Crops at the University of California, Riverside. Lyons had come to Australia to study the difference between chilling-resistant and chilling-sensitive plants. He believed that sensitivity to chilling was related in some way to an activity associated with the mitochondria.
Lyons and Raison devised methods of isolating mitochondria from these plants so that they could compare the rates of respiration of the different mitochondria at chilling temperatures. They found that over a range of 30 °C to 0 °C the respiration rate of mitochondria from resistant plants decreased by a factor of about 2 for each 10 °C decrease in temperature – a predictable result. However, with mitochondria from sensitive plants the results below 10 °C were quite unexpected. The decrease in respiration rate from 10 °C to 0 °C was much greater – in fact it decreased by a factor of 4.
Raison vividly recalls the day this result was first apparent. He took home the data collected during the day and that evening, with Lyons, began the drudgery of calculating the results. By 10 pm they had a clear picture which confirmed their belief. As John Raison later recalled:
I don't think you could describe the elation Jim and I went through at that moment. I think it is the height of any biologist's career to find that you can relate the behaviour of a whole organism to the behaviour of one tiny structure in its cells. To predict what might happen and then see, in the final analysis, that that is what did happen.
The two scientists toasted their success with glasses of Drambuie, then worked on until 3 o'clock in the morning, working out how this discovery could help explain the changes observed in plant tissues during chilling and why valuable crops like melons, tomatoes and avocados go mushy when stored at chilling temperatures.
Before Lyons returned to California, the two scientists extended their findings to animals. Mitochondria from the liver of (warm-blooded) rats showed the same response to lowered temperature as chilling-sensitive plants except that the sudden decrease in respiration rate occurred at 23 °C instead of 10 °C. Later, when Raison went to the University of California, Riverside, experiments with freshwater trout showed a direct similarity between the behaviour of mitochondria of cold-blooded animals and chilling-resistant plants – a constant decrease in respiration rate over the temperature range 30 °C to 0 °C.
From this point they began looking for an explanation for the unexpectedly rapid decline in the respiration rate of mitochondria from chilling-sensitive organisms. A study of lipids (fatty substances) in the mitochondrial membranes revealed that the lipids of chilling-resistant plants and cold-blooded animals remain semi-fluid at low temperatures. But, in chilling-sensitive plants and warm-blooded animals the lipids congeal at low temperatures – like butter in a refrigerator – causing a change in respiratory activity. The membrane walls become like an open mesh instead of a permeable plastic bag – energy production slows down, foreign substances leak in and the mitochondrial powerhouse collapses.
At the time the techniques to probe membrane structure were still in their infancy. Raison used electron spin resonance labelling to verify the physical changes in membrane lipids occurring in his experiments. These studies were done in collaboration with Alec Keith, a pioneer of the spin labelling method. The data were historically significant as they were made just after the fluid mosaic model of membrane structure had been proposed by SJ Singer (UC San Diego, La Jolla, California) and GL Nicolson (Salk Institute, La Jolla, California) in the 18 February 1972 issue of the prestigious journal Science. As a result membrane fluidity began to emerge as a hitherto unrecognised modulator of cell functions.
The work pioneered by John Raison was immediately recognised world-wide for its far reaching implications and was subsequently extended by him and his many collaborators to explain in molecular terms many of the interactions of temperature in relation to germination, photosynthesis, acclimation, torpor and the homeothermic status of heterothermic mammals.
With regard to the latter, further work in California showed that hibernating animals such as the golden-mantled ground squirrel have the ability to modify their membrane lipids before hibernation, so that they remain semi-fluid at low temperatures. Back in Australia by Raison showed that the same thing happens in the echidna.
John Raison and his fellow workers went on to identify chilling-resistant characteristics in plants, which will help plant breeders select chilling-resistant plants more easily and improve the keeping quality of their fruit. They also predicted that in time chemical or genetic means may be found for manipulating membrane lipids which may make it possible to extend the geographic range of many important food crops, to store sensitive fruit and vegetables for longer periods at much lower temperatures, and to extend considerably the storage time of vital human organs in transplant 'banks'.
- McKay A, 1976, 'Resisting the cold', Surprise and Enterprise, Fifty Years of Science for Australia, White F, Kimpton D (eds), CSIRO Publishing, pp.28.
- McMurchie T, 1991, 'John Raison - a tribute', Australian Society for Biochemistry and Molecular Biology Newsletter, May 1991, pp.20-21.