Cold acclimation in temperate, woody plants is a complex phenomenon that involves distinct changes in gene activity and protein synthesis. In previous research, a 60-kDa protein (PCA60), belonging to the dehydrin family of stress-related proteins, was identified in peach bark, and its corresponding gene (ppdhn1) was cloned and characterized. Presently, we report on the results of immunolocalization studies and in vitro cryoprotection assays. Seasonal collections of current-year stems were embedded in LR White or epoxy resin and sections of bark were probed with either a polyclonal antibody directed against a 15 amino acid sequence consensus region of dehydrins or a polyclonal antibody directed against partially-purified PCA60. In vitro cryoprotection assays utilized lactose dehydrogenase (LDH), a cold-labile enzyme. Immunolocalization at the light level indicated that the dehydrin was confined to the cytoplasm and absent in organelles. This localization was preliminarily confirmed at the ultrastructural level. LDH assays indicated cryoprotective activity in total protein extracts collected from winter bark tissues but completely absent in extracts of summer bark tissues. Preliminary LDH assays utilizing purified PCA60 also demonstrated cryoprotective activity. In general, the data further support a role for dehydrins in cold acclimation of woody plants.
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