Differences in freezing dynamics in the tip and base of wheat (Triticum aestivum L.) leaves result in a difference in cold hardiness

• Leaf tip burn is a freezing damage symptom in wheat, but its cause remains unclear. • Leaf tips are 3 K less cold-hardy than bases, despite similar mesophyll cell architecture. • Leaf tips froze faster, deviated more from equilibrium freezing and had less ice. • Raman imaging detected differences in cell wall chemistry between and within tissue types. • Lower cold hardiness was linked to less esterified cinnamic acids in cell walls and higher osmolarity. Leaf tip burn is a symptom of frost damage in wheat, though its cause remains unclear. Evidence suggests it is not caused by earlier ice formation but may occur during a third stage of freezing, likely due to differences in freezing dynamics between the leaf base and tip. We analyzed differences between leaf tips and bases of cold acclimated wheat, Triticum aestivum (cv. Shirley), with respect to leaf anatomy and microchemistry, cold hardiness, osmotic potential, spaces for apoplastic ice accumulation and freezing dynamics. Leaf tips were found to be 3 K less cold-hardy than the bases. Intracellular freezing was not observed; however, leaf tips froze faster, deviated more from ideal equilibrium freezing, and had a lower percentage of frozen water. Anatomical cell architecture did not explain differences in cold hardiness, but differences in osmolarity and in cell wall chemistry, particularly an increased esterification of cinnamic acids in the mesophyll at the base. Leaf tip burn in wheat is attributed to differences in cold hardiness along the leaf blade, resulting from developmental and physiological differences despite identical genetic backgrounds and growth conditions. This provides insights into the mechanisms responsible for increased cold hardiness.