Control of Ice Formation

To survive sub-freezing temperatures, organisms have developed a variety of mechanisms to maintain at least a minimum portion of their cellular liquids unfrozen. Variations in frost tolerance between acclimated and non-acclimated plants are at least partly explained by differences in the onset temperature of detrimental intracellular ice formation. Figure 4.17 shows the example of rye.

Whether crystallisation takes place inside or outside the cells depends on the cooling rate. When tissue is cooled slowly, water can move or leak out of the cells and crystallise outside the cell membranes in the extracellular spaces. This is a reversible process following the equilibrium between the water potentials of ice and of the cellular solutions. The water potential of ice is more negative than that of supercooled pure water.

Fig. 4.17. Formation of intracellular ice in protoplasts from frost-sensitive a and frost-hardened b rye leaves as a function of the cooling rate and the minimum temperature (x axis). Protoplasts from frost-sensitive leaves do not deposit plasma membrane material on the outer surface upon shrinking by extracellular ice formation. Intracellular ice formation is lethal for the cells. (Modified from Dowgert and Steponkus (1983))

The lower the sub-zero temperature is, the more negative the water potential of ice is and the more cellular water crystallises in the intercellular space. At a high cooling rate, there is not sufficient time for water export, and freezing takes place intracellularly, thereby killing the cells. In that case, ice crystals can grow through plasmodesmata and pits from cell to cell and rapidly expand to wider areas of the tissue.

For crystallisation, the liquid accumulating in the apoplastic space requires a nucleation trigger. Ice spreading from nearby xylem vessels could play such a role. In frost-sensitive dicots, ice spreads from only one nucleation event rapidly through the entire plant, while in graminoids, because of the polystele, each grass leaf requires separate nucleation. Thus, the shoot structure and vascular system are important for the spreading of ice and freezing damage (Hacker and Neuner 2007, 2008). Otherwise, so-called ice nucleation-active bacteria (INA bacteria; Box 4.4 in the substomatal cavity or on the surface of a leaf could trigger nucleation.

With potato and cauliflower, it has been shown that even water droplets on a leaf or a flower bud surface can catalyse ice formation inside the plant, and it takes only a few minutes for the entire plant to be frozen (Fuller and Wisniewski 1998). Ice nucleation activity has recently been reported in plant cell walls of bark tissue from blueberry stems, with outstanding activity (ice nucleation at -1.0 °C). This activity showed seasonal changes, peaking in November shortly before the onset of frost (Kishimoto et al. 2014). The mechanism underlying this INA principle is not yet known.