The <i>AtHDA6</i>‐<i>AtSK2</i> module promotes cold tolerance by enhancing shikimate metabolism and antioxidant activity

Low temperature is an environmental factor that significantly impairs the normal development of plants by limiting yield and quality. Although histone deacetylase HDA6 is involved in various biological processes, the specific molecular mechanisms underlying its response to low temperatures remain unexplored in Arabidopsis. In this study, we investigated the HDA6 expression pattern at low temperatures and discovered that cold stress-induced transcriptional activity increased the HDA6 protein level. Freezing experiments demonstrated that HDA6 functions as a positive regulator in response to low temperatures. The point mutant axe1-5 and the HDA6 CRISPR-edited knockout mutants hda6^CR-1 and hda6^CR-2 exhibited significantly increased sensitivity to low temperature, while the HDA6-GFP/axe1-5 complementation line successfully restored the cold-sensitive phenotype of the axe1-5 mutant. HDA6 interacted with and deacetylated shikimate kinase SK2. Furthermore, HDA6 enhanced SK2 protein stability under cold stress. The SK2-mediated shikimate metabolic pathway is crucial for the synthesis of aromatic amino acids, which are essential antioxidant precursors. Metabolomics analysis showed that the hda6 mutant metabolites that decreased significantly under cold stress were primarily concentrated in the amino acid synthetic pathway. Additionally, the hda6 and sk2 mutants accumulated higher levels of superoxide anion but lower levels of antioxidant substances under cold stress, suggesting that HDA6 may enhance shikimate metabolism, downstream amino acid synthesis, and antioxidant accumulation by stabilizing SK2, thereby improving cold tolerance. This study elucidated the molecular mechanism by which HDA6 positively responds to low-temperature stress and identified the antifreeze genes HDA6 and SK2. This study offers valuable genetic resources and theoretical support for breeding cold-resistant varieties and improving crop yield.