Abstract
Populus simonii × P. nigra is a valuable model for studying woody plant responses to saline-alkaline stress, however the molecular mechanisms underlying its adaptation remain poorly understood. Here, integrated transcriptomic and metabolomic analyses were used to examine the responses in leaves to low (50 mmol/L) and high (200 mmol/L) alkaline salt stress over 7 and 14 days. Physiological assessments revealed that high-concentration stress inhibited growth, induced oxidative damage, elevated malondialdehyde (MDA) content, enhanced antioxidant enzyme activity (SOD, POD, CAT), and promoted osmoregulatory compound accumulation (soluble sugars, proline, free amino acids). RNA sequencing revealed extensive transcriptional reprogramming, with more pronounced changes under higher stress and longer exposure. KEGG enrichment highlighted pathways such as plant hormone signal transduction starch/sucrose metabolism, and flavonoid and phenylpropanoid biosynthesis. Metabolomic profiling detected 4257 metabolites, with notable increases in amino acids, organic acids, sugars, and flavonoids. Integrated analyses revealed coordinated regulation of genes and metabolites in phenylalanine, tyrosine, and tryptophan biosynthesis, as well as hormone signaling. Weighted gene co-expression network analysis (WGCNA) identified modules correlated with physiological traits, enriched in stress-responsive transcription factors (MYB, AP2/ERF, NAC, WRKY) and metabolic pathways. Key genes in starch/sucrose degradation, and raffinose metabolism were upregulated, whereas phenylpropanoid metabolism-related genes were suppressed. Collectively, these findings provide insight into the adaptive mechanisms of P. simonii × P. nigra to saline-alkaline stress, highlighting regulatory genes and pathways that contribute to tolerance and offering potential targets for enhancing poplar resilience in alkaline salt environments.