Genome-Wide Profiling of Cold-Responsive Genes in Zelkova schneideriana Under Chilling Stress

Zelkova schneideriana Hand.-Mazz. is an ecologically and economically valuable tree species in China, but its sensitivity to low temperature restricts its use in colder regions. To identify genes and pathways associated with chilling responses, we combined physiological screening, full-length transcriptome construction, Illumina RNA-seq, and time-ordered gene co-expression network analysis (TO-GCN). Fifty seedling lines were evaluated after 7 d at 4 °C, and the line with the lowest relative electrolyte leakage (REL) and malondialdehyde (MDA) content was selected for transcriptome profiling at 0, 3, and 6 d of chilling treatment. RNA-seq detected 5875 differentially expressed genes. Most expression changes occurred by day 3 and were largely maintained on day 6. Pathway reconstruction showed contrasting dynamics between antioxidant-related systems: Flavonoid and anthocyanin biosynthesis were progressively activated, whereas glutathione-related genes, particularly glutathione S-transferase family members, were broadly repressed. TO-GCN highlighted ZeF3H (Ze_transcript_14112) as a late-stage hub associated with anthocyanin biosynthesis and ZeDRT102 (Ze_transcript_44427) as an early-stage hub linked to DNA damage repair and glutathione-associated genes. These results indicate that the chilling response in Z. schneideriana involves activation of flavonoid-based protection together with weakened glutathione-mediated redox regulation, providing candidate genes for functional validation and cold-resistance breeding. These insights provide a critical foundation for molecular breeding strategies that will ultimately facilitate the successful introduction and stable cultivation of this valuable species in northern temperate climates.