Physiological and transcriptome analyses for revealing the rooting mechanism of Mussaenda anomala, a species with an extremely small population

• Histological studies of cells revealed that the ARs of Mussaenda anomala originate from epidermis cells. • Through integrating multi-hormone-key enzyme activity-metabolite accumulation-transcriptional dynamics, the mechanism of ARs formation in M. anomala was analyzed. • SOD continues to rise, indirectly promoting IAA synthesis; CAT, POD, ACC, JA, and SA jointly drive rooting through redox and hormone interactions; low levels of tZR, GA1, and ABA are conducive to root formation and elongation. • This study identified three modules and 11 key genes, and clarified their positive/negative regulative roles in ARs development of M. anomala . This study investigates the mechanism of adventitious roots (ARs) formation in Mussaenda anomala cuttings. We integrated anatomical structure observation, physiological index measurement, and transcriptome analysis at four stages: initial cutting (D1), ARs primordia induction (D2), ARs formation (D3), and ARs elongation (D4). The results revealed that root primordia originate from epidermis cells; SOD activity continuously increased, indirectly promoting IAA biosynthesis and ARs development. Meanwhile, CAT, POD, ACC, JA, and SA collectively drive ARs root formation through redox balance and hormone signaling. Additionally, PPO and IAAO synergistically participated in wound healing and IAA homeostasis, leading to the consumption of substantial soluble sugars and proteins. Low levels of tZR, GA1, and ABA further facilitated ARs development. Furthermore, transcriptome analysis identified key gene modules: the hub genes in the green module ( NRT1/PTR 7.3-like, NRT1/PTR 5.10-like, CYP 93A3-like, CYPRO4-like ), and brown module ( TIFY 6B-like, CRF4, CYP94A1-like, AGP 27-like/ADGP ) positively regulated ARs formation; whereas the hub genes in the turquoise module ( ADH-like, ZFP16-like, MYB86-like ) acted as negative regulators. These results clarify the regulatory network underlying ARs development in M. anomala , supporting its vegetative propagation and germplasm conservation.