Metabolic Dynamics and Modular Regulatory Mechanisms of Leaf Abscission in Cyclocarya paliurus Stem Segments In Vitro

Leaf abscission is extremely severe during the Cyclocarya paliurus stem segment formation in vitro culture, and stem segment development is hindered after leaf abscission. To explore the dynamic regulatory mechanisms of metabolites in the leaf abscission process of C. paliurus, the emerged leaves of C. paliurus stem segments were cultured for 22 days (T0) in vitro; leaves at 27 days (T1) and leaves that had fallen after ≥ 32 days (T2) were used as materials for analysis of the types and contents of metabolites by liquid chromatography-tandem mass spectrometry (LC-MS/MS). A total of 2160 differentially accumulated metabolites (DAMs) were obtained across the three collected time points. KEGG enrichment analysis showed significant enrichments in both flavonoid biosynthesis and C5-branched dibasic acid metabolism. Based on co-expression network analysis, four modules significantly associated with abscission were identified. The turquoise module genes (CAT1-like, RAX2-like MYB, E2 4-like, and RBOH) promote flavonoid metabolite biosynthesis and synergistically drive abscission through oxidative stress and cell wall degradation. In contrast, the yellow module genes (14-3-3, MAPKK, ERF4, and ERF2) tend to maintain C5-branched dibasic acid metabolism and auxin transport homeostasis, while suppressing the ABA/ethylene-driven senescence pathway. The green module genes (Aux/IAA13-like, OPCL1) and blue module genes (HCT, CALDH) weaken auxin signaling and cell wall structural stability. These four modules work synergistically to collectively promote the leaf abscission process in C. paliurus. This study provides novel insights into the molecular regulatory mechanisms underlying leaf abscission in stem segments of C. paliurus cultured in vitro.