Molecular mechanisms and energetic costs of recovery from freezing in the polyextremophile midge, Belgica antarctica

Extracellular freezing challenges insects in several ways, including mechanical damage from ice crystals, exposure to stressful levels of cold and cellular dehydration. While stress-response mechanisms activated during and immediately after freezing are well studied, less is known about how freeze-tolerant organisms recover from freezing in the longer term and whether this recovery carries energetic costs. Here, we tracked changes in gene expression and energy stores over the course of 15 days following extracellular freezing using the world's southernmost insect, Belgica antarctica (Diptera: Chironomidae), as a study system. We found that B. antarctica employed a coordinated ‘emergency’ stress-response system during early recovery (0–1 day), which includes the upregulation of genes involved in inhibiting cell death (e.g. Bcl-2, IAP-1) alongside mechanisms involved in damage repair and clearance of cell debris (e.g. heat shock proteins, autophagy, proteasome). Concomitantly, genes involved in ecdysone biosynthesis and juvenile hormone degradation were downregulated through the 1st day of recovery, and several genes involved in cuticle development were downregulated between the 3rd and 15th day of recovery, suggesting that homeostasis may not have been completely restored for 2 weeks after B. antarctica had thawed. These physiological changes did not lead to a detectable depletion of energy stores, indicating that this emergency response system does not incur significant energy drain. These results suggest that evoking a coordinated stress response with minimal energy usage may be crucial for B. antarctica to persist in cold environments, and that blocking apoptosis and pausing development could provide sufficient time for freezing injury to be fully addressed.