Autophagy and energy metabolism drive molting regulation in Macrobrachium rosenbergii: Insights from histological, physiological, and transcriptomic analyses

Molting is a critical event throughout the life cycle of crustaceans; however, the regulatory mechanisms underlying molting in Macrobrachium rosenbergii remain poorly understood. In this study, we employed transmission electron microscopy, molecular biology techniques, and transcriptome sequencing to systematically investigate the molting regulation in M. rosenbergii. The results revealed that the exoskeleton cuticle primarily consists of the epicuticle, exocuticle, and endocuticle. The cuticle underwent periodic structural remodeling during the molting cycle, with its thickness initially increasing and subsequently decreasing. Additionally, the hemolymph levels of 20-hydroxyecdysone (20E), crustacean hyperglycemic hormone (CHH), and the energy molecule ATP were significantly elevated during the molt and post-molt stages compared to the inter-molt and pre-molt stages, and melatonin (MT) content was significantly higher in post-molt stage than the other stages (p < 0.05). Transcriptome analysis revealed that differentially expressed genes (DEGs) were significantly enriched in the lysosomal pathway. Notably, the expression levels of key DEGs (e.g., Lmp2l, Mfsd6l, ATP13a3, Pipoxl) were significantly correlated with ecdysteroid levels (p < 0.05). Specifically, 20E levels were positively correlated with the expression of Mfsd6l and ATP13a3 during the pre-molt stage, while CHH levels were positively correlated with Pipoxl expression in the post-molt stage (p < 0.05). In the post-molt stage, interaction network analysis indicated that highly expressed genes were primarily enriched in the lysosome, steroid hormone biosynthesis, retinol metabolism, AMPK, and FoxO pathways. Collectively, these findings highlight the central roles of autophagy (lysosomal pathway) and energy metabolism (AMPK/FoxO pathways) in molting regulation, providing essential data for advancing the understanding of molting physiology and improving culture management strategies.