Detection of polypeptides associated with the cold acclimation process in spinach

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Abstract
Exposure of spinach (Spinacia oleracea) seedlings to 5 °C for several days has previously been shown to induce a greater tolerance to the stresses of extracellular freezing. Associated with this response to low temperature, termed cold acclimation, was a subtle shift in protein synthesis and altered polypeptide composition. Two‐dimensional gel electrophoresis was used to study the changes in spinach leaf tissue protein synthesis in an effort to identify polypeptides that may play a central role in the induction of greater freezing tolerance. Through a combination of silver staining, in vivo labeling, and in vitro translation of mRNAs, we identified several high molecular weight polypeptides whose synthesis and presence in spinach leaf tissue were highly correlated with freezing tolerance. Synthesis of these polypeptides was elevated or induced during cold acclimation when freezing tolerance increases, but was rapidly reduced or halted during deacclimation when freezing tolerance declines. The close association of the synthesis of these polypeptides with the induction and loss of freezing tolerance suggested that they could play a role in cold tolerance mechanisms of spinach.