Transcriptome and lipidome analyses reveal membrane lipid remodeling in root tissues of Cannabis sativa L. exposed to cold stress.

Yan B.; Chang C.; Sui Y.; Zhao X.; Zheng N.; Fang Y.; Zhang Y.; Zhang M.; Zhang L. · Plant physiology and biochemistry : PPB · 2026

**Background & Methods** This study examined cold stress responses in Cannabis sativa L. (hemp cultivar "Longdama 9") root tissues using integrated phenotypic, physiological, biochemical, lipidomic, and transcriptomic analyses. Roots were exposed to 4°C for 7 days, with comprehensive molecular and cellular assessments performed to construct a regulatory network model of cold tolerance mechanisms. **Key Findings** • **Structural and Physiological Damage**: Cold stress significantly inhibited root growth, with 24% reduction in root volume and 28% decrease in fresh weight. Membrane damage was substantial, evidenced by 2.9-fold increase in electrical conductivity and 3.8-fold elevation in malondialdehyde content, indicating lipid peroxidation. • **Adaptive Metabolic Responses**: Hemp roots activated multiple protective mechanisms including accumulation of osmotic regulators (soluble sugars, proteins, proline) and increased antioxidant enzyme activity (SOD, POD, CAT). These coordinated responses preserved cellular homeostasis under cold conditions. • **Membrane Lipid Remodeling**: Lipidomic analysis revealed dramatic lipid composition changes, including 25% reduction in phosphatidylcholine, 26% increase in phosphatidic acid, 27% increase in digalactosyldiacylglycerol, 46% decrease in monogalactosyldiacylglycerol, and 69% increase in triacylglycerol. Critical lipid ratios (PC/PE and [PI+PC]/[PA+PE]) decreased while DGDG/MGDG ratio increased. • **Transcriptional Regulation**: Transcriptomic analysis identified 5,786 differentially expressed genes, including 730 lipid metabolism-associated genes. Upregulated genes included those encoding GPAT, LPAT, DGAT, PLDα, and fatty acid desaturases (FAD4, FAD6, FAD8). The zinc finger transcription factor CsZincF was demonstrated to directly activate CsDGAT2 transcription through promoter binding assays. **Dosage & Administration** Not reported. **Safety & Adverse Effects** Not reported. **Evidence Quality** This is a controlled laboratory study with comprehensive multi-omics integration, providing strong mechanistic evidence. However, limitations include single cultivar testing, controlled greenhouse conditions not reflecting field environments, and reliance on correlation rather than causation for gene-phenotype relationships. Results require validation across diverse hemp cultivars and natural cold stress conditions. The Y1H and dual-luciferase assays provide solid functional evidence for specific transcriptional regulation but warrant additional in vivo validation.

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