Physical, chemical, and physiological characterization of the reactions preceding chilling injury–induced cell membrane damage in cucumber

Abstract

Chilling injury in cucumber is commonly thought to begin with cell membrane damage caused by reactive oxygen species, which results in ion leakage from the tissue. However, additional short-term storage at ambient temperature after cold storage can accelerate the appearance of damage, and the different types of vegetables exhibit distinct symptoms. We hypothesized that cell membrane damage is a secondary reaction, and that preceding responses in the tissue lead to irreversible changes. To explore these responses in cucumber tissue during cold storage, we employed several techniques, including ion leakage measurements, bio-electrochemical impedance spectroscopy, X-ray computed tomography, nuclear magnetic resonance–based metabolite profiling, and transcriptome analysis. Our time-course comparison of different storage conditions (2 °С, 5 °С, 10 °С, and 15 °С for 0–14 d) indicated that several responses preceded detectable ion leakage–associated cell membrane damage. Specifically, the cold-stored samples exhibited abnormal electrolyte distribution and marked changes in metabolite levels and gene expression related to the tricarboxylic acid (TCA) cycle, which is critical for generating energy for homeostasis. Moreover, samples stored at 2 °С followed by 15 °С showed earlier initiation of tissue responses in metabolite and gene expression profiles, suggesting the occurrence of irreversible changes, possibly in the TCA cycle. These findings suggest that the critical response to chilling injury may occur prior to cell membrane damage, likely due to metabolic imbalances in the TCA cycle.