Redesigning Crops for Increased Tolerance to Freezing Stress

Abstract

The process of cold acclimation is a complex developmental phenomenon that invokes the orchestration of many different processes, including hormonal responses to environmental cues, altered gene activity, the synthesis of new gene products, and alterations that influence virtually every facet of cellular metabolism. Although seemingly disparate, many of these metabolic changes ultimately contribute to the increased cryostability of cellular membranes, destabilisation of which is the primary cause of freezing injury. Therefore, to provide a rational integration of the many different facets of the cold acclimation process, it is necessary to first identify the freeze/thaw-induced lesions in cellular membranes and to elucidate the biophysical mechanisms and biochemical alterations responsible for their occurrence. Although all cellular membranes are vulnerable to freeze-induced destabilisation, maintenance of the structural integrity of the plasma membrane is a prerequisite for survival because of the central role that it plays during a freeze-thaw cycle. Cryomicroscopy studies of isolated protoplasts complemented with electron microscopy studies of freeze- induced ultrastructural changes are yielding a comprehensive analysis of the phenomenology of freezing injury and the identification of specific ‘lesions’ in the plasma membrane, which vary depending on the stage of acclimation and the nadir temperature to which the protoplasts are cooled. Of the lesions identified to date (expansion-induced lysis, lamellar-to-hexagonal II phase transitions, and the fracture-jump lesion), all are a consequence of freeze- induced dehydration; however, whereas expansion-induced lysis is the result of cellular dehydration and the large osmotic excursions incurred during a freeze-thaw cycle, lamellar-to-hexagonal II phase transitions and the fracture-jump lesion are consequences of the removal of water that is closely associated with cellular membranes. These studies, together with a detailed analysis of the plasma membrane lipid composition at the molecular species level and procedures to alter selectively the lipid composition of the plasma membrane have provided a foundation for mechanistic studies to establish directly the relationships between alterations in the plasma membrane lipid composition and its increased cryostability after cold acclimation. In order to determine if genotypic diversity in freezing tolerance is associated with genotypic differences in membrane lipid composition, we are extending these studies to other cereals (e.g., oat and barley), which are less freezing tolerant than winter rye. These studies reveal that the freeze-induced membrane lesions in a spring oat cultivar (Ogle) are qualitatively similar to those observed in rye, but occur at substantially higher temperatures. Moreover, the lipid composition of the plasma membrane isolated from leaves of the spring oat is vastly different from that of winter rye leaves. These studies suggest that strategies for improving the freezing tolerance of cereals should include techniques to modify membrane lipid composition.