Responses to Ice Formation and Reasons of Frost Injury in Potato Leaves

Abstract

Potato leaves are ice-tolerant but are frost-damaged at −3 °C. Freezing occurs in two steps, a first non-destructive freezing event and a second independent lethal event. Localization of ice, and whether cells freeze-dehydrate after the first freezing event remains unknown. The cause of frost damage during the second freezing event lacks experimental evidence. Cytological responses of mesophyll cells were examined during ice formation using cryo-microscopic techniques after high-pressure freeze-fixation and freeze-substitution. CO2 gas exchange on frozen leaves revealed functional responses, but also frost damage. After the first freezing event, gas exchange was uninterrupted. Consequently, intercellular spaces are free of ice, and ice may be restricted to xylem vessels. The cellular shape of the mesophyll cells was unchanged, cells did not freeze-dehydrate but were supercooled. When thawed after the first freezing event, leaves were initially photoinhibited but regained photosynthesis. During the second freezing event, cells froze intracellularly, and some palisade parenchyma cells remained intact for a prolonged time. Intracellular ice caused complete destruction of cells, and chloroplasts became invisible at the light microscopic level. When thawed after the second freezing, leaves were unable to regain photosynthesis. Consequently, freezing avoidance is the only viable strategy for potatoes to survive frost.