Destabilization of the plasma membrane, which is a primary cause of freezing injury, is a consequence of freeze-induced osmotic stresses and cell dehydration. However, the mechanism of injury depends on the magnitude of the osmotic stress and the extent of cell dehydration. Over the range of 0 to — 5 °C, destabilization of the plasma membrane in protoplasts isolated from non-acclimated rye leaves is a result of osmotic excursions, because freeze-induced osmotic contraction results in endocytotic vesiculation of the plasma membrane and sufficiently large area reductions are irreversible. At lower temperatures, the protoplasts are subjected to extremely large osmotic pressures ( — 12 MPa at —10 °C), and there are several changes in the ultrastructure of the plasma membrane, including the formation of aparticulate domains and lamellar- to hexagonaln-phase transitions. These changes, which are manifestations of demixing of the membrane components, are predicted by a theory of bilayer interactions at low levels of hydration. During cold acclimation, the cryobehaviour of the plasma membrane is altered; osmotic contraction results in the reversible formation of exocytotic extrusions and the propensity for dehydration-induced demixing and lamellar- to hexagonaln-phase transitions is decreased. In both cases, the differential behaviour is also observed in liposomes prepared from plasma membrane lipids isolated from non-acclimated and cold-acclimated leaves. However, as no lipid species are unique to the plasma membrane of either non-acclimated or cold-acclimated leaves, the differential behaviour is caused by altered lipid-lipid interactions because of different proportions of the lipid species. Hence the behaviour of the plasma membrane can be altered by using a protoplast-liposome fusion procedure to selectively modify the lipid composition of the plasma membrane. These studies provide direct evidence that the increased cryostability of the plasma membrane is a consequence of alterations in its lipid composition.
Behavior of the Plasma Membrane of Isolated Protoplasts during a Freeze-Thaw Cycle