Understanding the Regulation of Estivation in a Freshwater Snail through iTRAQ-Based Comparative Proteomics

The apple snail Pomacea canaliculata is a freshwater gastropod with a remarkable ability to withstand seasonal or unpredictable dry conditions by entering estivation. Studies of P. canaliculata using conventional biochemical and the individual gene approaches have revealed the expressional changes of several enzymes and antioxidative genes in response to estivation and arousal. In this study, we applied iTRAQ-coupled two-dimensional LC-MS/MS to identify and quantify the global protein expression during the estivation and arousal of P. canaliculata. A total of 1040 proteins were identified, among which 701 proteins were quantified and compared across four treatments (i.e., control, active snails; short-term estivation, 3 days of exposure to air; prolonged estivation, 30 days of exposure to air; and arousal, 6 h after resubmergence in water) revealing 53 differentially expressed proteins. A comparison of protein expression profiles across treatments indicated that the proteome of this species was very insensitive to initial estivation, with only 9 proteins differentially expressed as compared with the control. Among the 9 proteins, the up-regulations of two immune related proteins indicated the initial immune response to the detection of stress cues. Prolonged estivation resulted in many more differentially expressed proteins (47 compared with short-term estivation treatment), among which 16 were down-regulated and 31 were up-regulated. These differentially expressed proteins have provided the first global picture of a shift in energy usage from glucose to lipid, prevention of protein degradation and elevation of oxidative defense, and production of purine for uric acid production to remove toxic ammonia during prolonged estivation in a freshwater snail. From prolonged estivation to arousal, only 6 proteins changed their expression level, indicating that access to water and food alone is not a necessary condition to reactivate whole-sale protein expression. A comparison with hibernation and diapause revealed many similar molecular mechanisms of hypometabolic regulation across the animal kingdom.