Translational regulation in the anoxic turtle, Trachemys scripta elegans

Abstract

The red-eared slider turtle (Trachemys scripta elegans), has developed remarkable adaptive mechanisms for coping with decreased oxygen availability during winter when lakes and ponds become covered with ice. Strategies for enduring anoxia tolerance include an increase in fermentable fuel reserves to support anaerobic glycolysis, the buffering of end products to minimize acidosis, altered expression in crucial survival genes, and strong metabolic rate suppression to minimize ATP-expensive metabolic processes such as protein synthesis. The mammalian target of rapamycin (mTOR) is at the center of the insulin-signaling pathway that regulates protein translation. The present study analyzed the responses of the mTOR signaling pathway to 5 (5H) or 20 h (20H) of anoxic submergence in liver and skeletal muscle of T. scripta elegans with a particular focus on regulatory changes in the phosphorylation states of targets. The data showed that phosphorylation of multiple mTOR targets was suppressed in skeletal muscle, but activated in the liver. Phosphorylated mTOR showed no change in skeletal muscle but had increased by approximately 4.5-fold in the liver after 20H of anoxia. The phosphorylation states of upstream positive regulators of mTOR (p-PDK-1, p-AKT, and protein levels of GβL), the relative levels of dephosphorylated active PTEN, as well as phosphorylation state of negative regulators (TSC2, p-PRAS40) were generally found to be differentially regulated in skeletal muscle and in liver. Downstream targets of mTOR (p-p70 S6K, p-S6, PABP, p-4E-BP1, and p-eIF4E) were generally unchanged in skeletal muscle but upregulated in most targets in liver. These findings indicate that protein synthesis is enhanced in the liver and suggests an increase in the synthesis of crucial proteins required for anoxic survival.

MicroRNAs regulate survival in oxygen-deprived environments

“…The molecular mechanisms that underlie anoxia tolerance adaptations include: post-translational modifications, epigenetic regulation, transcription factor regulation, the accumulation of organic osmolytes, and the post-transcriptional regulation of small regulatory non-coding RNAs called microRNAs (miRNAs) (Szereszewski and Storey, 2018;Wijenayake et al, 2018;Biggar and Storey, 2017;Green and Storey, 2016;Kornfeld et al, 2012;Krivoruchko and Storey, 2010;Muir et al, 2007). miRNAs are highly conserved sequences of non-coding RNA approximately 22 nucleotides long that have been identified as critical regulators of a diverse range of cellular functions through their ability to suppress translation (Lund et al, 2004;Christopher et al, 2016).…”

mTOR Signaling in Metabolic Stress Adaptation

“…In the past decade, we and others have identified mTORC1 as a key regulator governing animal survival under various environment-induced situations of metabolic rate depression. These have included hibernation in mammals [ 15 , 16 ], anoxia-tolerance of submerged turtles [ 17 ], hypoxia-tolerance of naked mole rats [ 18 ], and dehydration-induced estivation in frogs [ 19 ]. A surprising finding across these studies is that mTORC1 can be both suppressed and activated during periods of metabolic depression, suggesting that its functional role is stress-type and species-dependent.…”

“…One study measuring de novo protein synthesis in red-eared sliders indicated that the rate of radiolabeled 35 S-methionine incorporation was not substantially reduced after 20 hours of anoxia exposure, and 35 S-methionine was actually elevated in select tissues including the liver and heart [ 69 ]. Biochemical studies have shown that levels of p-Akt (Ser-473) were elevated in the liver and muscle of red-eared slider turtles during anoxia, meanwhile, p-mTOR was maintained at a similar level compared to controls in the liver but increased by ~4.5-fold in the skeletal muscle [ 17 ]. Consistent with mTORC1 being activated during oxygen deprivation, levels of p-S6 protein and p-4E-BP1 were also highly increased in turtles after 20 hours of anoxia exposure [ 17 ].…”

“…Biochemical studies have shown that levels of p-Akt (Ser-473) were elevated in the liver and muscle of red-eared slider turtles during anoxia, meanwhile, p-mTOR was maintained at a similar level compared to controls in the liver but increased by ~4.5-fold in the skeletal muscle [ 17 ]. Consistent with mTORC1 being activated during oxygen deprivation, levels of p-S6 protein and p-4E-BP1 were also highly increased in turtles after 20 hours of anoxia exposure [ 17 ].…”

“…Phosphorylation (P) in green indicates the activation effect while phosphorylation in red indicates inhibitory effect. Data are summarized from the following studies: [ 15 , 16 , 17 , 18 , 19 , 46 , 51 , 52 ].…”