Induction of Hibernation and Changes in Physiological and Metabolic Indices in Pelodiscus sinensis

SIMPLE SUMMARY: Hibernating animals reduce their metabolic rate through a variety of transcriptional, translational, and post-translational regulation methods to survive in extreme environments with an overall reduction in their body’s activity levels. Histone methylation is a mechanism that regulates gene expression at the transcriptional level, and specific histone methylation and demethylation modifications can either inactivate or activate gene transcription. In this study, we focused on the hibernation of Pelodiscus sinensis (P. sinensis) as an experimental model and studied the changes in the physiological metabolism index, histones, and related genes on growth and development under conditions of artificially induced hibernation. Although this paper lacks further evidence that P. sinensis has been induced into a deep hibernation state, cold torpor is perhaps a more accurate description. The changes in physiological indexes, transcription level, mRNA relative expression, protein localization, relative protein expression, and enzyme activity associated with histone and histone (demethylation) genes in various tissues were analyzed under hibernating and normal conditions. The results provide insight into the adaptative mechanism of P. sinensis to its environment. ABSTRACT: Pelodiscus sinensis (P. sinensis) is a commonly cultivated turtle species with a habit of hibernation. To study the changes in histone expression and methylation of P. sinensis during hibernation induction, a model was established by artificial induction. Physiological and metabolic indices were measured, and the expression and localization of histone (H1, H2A, H2B, H3, and H4) and methylation-related genes (ASH2L, KMT2A, KMT2E, KDM1A, KDM1B, and KDM5A) were measured by quantitative PCR, immunohistochemistry, and Western blot analysis. The results indicated that the metabolism, antioxidation index, and relative expression of histone methyltransferase were significantly decreased (p < 0.05), whereas the activity and expression of histone demethyltransferase were significantly increased (p < 0.05). Although our results showed significant changes in physiological and gene expression after hibernation induction, we could not confirm that P. sinensis entered deep hibernation. Therefore, for the state after cooling-induced hibernation, cold torpor might be a more accurate description. The results indicate that the P. sinensis can enter cold torpor through artificial induction, and the expression of histones may promote gene transcription. Unlike histones expressed under normal conditions, histone methylation may activate gene transcription during hibernation induction. Western blot analysis revealed that the ASH2L and KDM5A proteins were differentially expressed in the testis at different months (p < 0.05), which may perform a role in regulating gene transcription. The immunohistochemical localization of ASH2L and KDM5A in spermatogonia and spermatozoa suggests that ASH2L and KDM5A may perform a role in mitosis and meiosis. In conclusion, this study is the first to report changes in histone-related genes in reptiles, which provides insight for further studies on the physiological metabolism and histone methylation regulation of P. sinensis during the hibernation induction and hibernation period.