Role of Glucose-induced Transcription Factor Signalling and Mitochondrial Epigenetics in Stress Tolerant Wood Frog, Rana sylvatica

The freeze-tolerant wood frogs, Rana sylvatica are one of only a few vertebrate species in the animal kingdom, which are extensively studied to understand vertebrate freeze tolerance. They undergo whole-body freezing during winter and become ice solid with no heartbeat, brain activity and blood flow but amazingly come back to life during spring unharmed without any major changes in their body. Freeze survival is challenging, with wood frogs facing ischemia due to freezing of blood, dehydration via cell volume reductions due to loss of 60-70% of total body water into extracellular space as well as hyperglycemia, producing a huge amount of glucose as a cryoprotectant. Interestingly, wood frogs can also tolerate these stresses independent of freezing. Also, winter survival by wood frogs is associated with a metabolic reorganization to reduce their energy demands to a bare minimum by globally suppressing energy-expensive pathways and selectively regulating genes to prioritize available energy use for pro-survival pathways. This thesis examined the effects of freezing and dehydration-induced hyperglycemic response in selectively inducing transcription factor MondoA in regulating glucose-induced transcription and activating an adaptive transcriptional response to induce stress response via inflammasome activation, mitochondrial dysfunction and mitochondrial epigenetics. The current findings establish MondoA in guiding an adaptive transcriptional response to activate genes regulating glucose homeostasis and circadian rhythm in a tissue-specific manner in the liver during the freeze/thaw cycle. Also, the role of TXNIP (downstream to MondoA) and its PTMs, in activating inflammasome via NLRP-3 in stress-specific way during freezing was shown. Moreover, the higher mitochondrial presence of TXNIP did not correlate to protein expression of its downstream targets in inducing mitochondrial dysfunction in any of the stresses, which were attributed to its low/weak binding to TRX-2. Investigating the role of mitochondrial methylation suggests its tissue-specific regulation in the liver and potential role in maintaining a tight regulation of mitochondrial transcriptional and gene expression response. Altogether, findings from this thesis demonstrate that a highly synchronized and intricate control via multiple levels of regulation is present in activating mechanisms that are involved in maintaining cellular milieu during stress in wood frogs.