Transcriptome and DNA methylation responses in the liver of Megalobrama amblycephala under hypoxia and heat stress

Global warming and aquaculture expansion are raising water temperatures and decreasing dissolved oxygen (DO) in freshwater environments, threatening fish survival. Revealing the molecular mechanisms of fish response to heat and/or hypoxia is vital for the development of sustainable aquaculture and genetic breeding strategies. This study examined the molecular adaptations of Megalobrama amblycephala, an important economic fish in China, to environmental stresses. Liver samples from fish subjected to hypoxia (LO), heat (HT), and their combination (HL) were analyzed using transcriptome and whole-genome methylation sequencing. Results indicated that environmental stresses significantly altered gene expression and DNA methylation levels in M. amblycephala, suggesting a molecular basis for enhanced environmental adaptability through metabolic regulation. Comparative analysis showed distinct differences in differentially expressed genes (DEGs) across the three stress groups, with the highest number occurring in the NC vs HL group, indicating a stronger transcriptional response. PPI analysis revealed significant alterations in metabolic pathway gene expression. Examination of the PPI network constructed from DEGs common to all stress groups pinpointed core metabolic regulators, such as the upregulated hk1 and aldoaa and the downregulated gck and fasn, highlighting a coordinated cellular metabolic adaptation to environmental stresses. DNA methylation analysis revealed CG-type methylation as the predominant pattern. The HL group exhibited elevated CHH and CHG methylation compared to other groups. Integration of DMR-promoter genes and DEGs yielded 12, 8, and 29 overlapping genes in the NC vs LO, NC vs HT, and NC vs HL groups, respectively. KEGG analysis of negatively regulated overlapping genes showed significantly enriched pathways: TGF-beta signaling pathway and endocytosis in the NC vs LO group; galactose metabolism and amino sugar and nucleotide sugar metabolism in the NC vs HT group; and other glycan degradation and various types of N-glycan biosynthesis in the NC vs HL group. These findings indicated that the adaptation of M. amblycephala to environmental changes is driven by coordinated changes in specific gene methylation and transcription levels, thereby fine-tuning physiological processes including intracellular signaling, glucose and glycoconjugate metabolism. These findings provide theoretical support for optimizing cultivation strategies of M. amblycephala.