Physiological Responses to Thermal Stress in the Liver of Gymnocypris eckloni Revealed by Multi-Omics

Climate-change-induced thermal stress poses a significant threat to cold-adapted aquatic species, particularly fish endemic to high-altitude ecosystems such as <i>Gymnocypris eckloni</i>, which is native to the Qinghai-Tibetan Plateau. To elucidate the molecular and metabolic mechanisms underlying their response to elevated temperatures, we integrated RNA-seq, miRNA-seq, and LC-MS-based metabolomic analyses of liver tissue from fish exposed to chronic thermal stress (HT) versus control (CT) conditions. Although no significant differences were observed in growth parameters, histopathological examination revealed structural damage under heat stress. Transcriptomic analysis identified widespread dysregulation of genes involved in energy metabolism, with significant downregulation of pathways related to amino acid, fatty acid, glucose, and oxidative phosphorylation. In contrast, upregulated DEGs were enriched in N-glycan biosynthesis, protein processing in the endoplasmic reticulum, and phagosome. Concomitant miRNA profiling revealed differentially expressed miRNAs, including miR-196a-5p, miR-132-3p, and miR-181b-5p, which were predicted to regulate key metabolic genes such as <i>ugt1a1</i>, <i>pepck</i>, and <i>calr</i>. Metabolomic analysis further demonstrated significant alterations in metabolic profiles, with glutathione metabolism, tryptophan metabolism, steroid hormone biosynthesis, and pyruvate metabolism emerging as central pathways in the heat stress response. Integrated multi-omics analysis confirmed coordinated regulation of these pathways, highlighting the critical role of glutathione and tryptophan, as well as disruptions in purine and energy metabolism. The DEMiR-DEG-DEM networks involving miR-196a-5p-<i>pepck</i>-PEP, miR-133a-3p-<i>gne</i>-UDP-GlcNAc, and miR-132-3p-<i>ugt1a1</i>-Bilirubin may play an important role in thermal stress. This study provided a new perspective on the molecular, regulatory, and metabolic adaptations of <i>Gymnocypris eckloni</i> to thermal stress, identifying potential biomarkers and regulatory networks that may inform conservation strategies for cold-water fish under global warming.