Activation of antioxidant defenses in response to freezing in freeze-tolerant painted turtle hatchlings.
BACKGROUND
Hatchlings of the painted turtle, Chrysemys picta marginata can endure long-term freezing of their extracellular body fluids. We hypothesized that freezing survival would include adaptive up-regulation of antioxidant defenses to deal with ischemia-reperfusion injuries associated with the freeze-thaw cycle. A number of antioxidant enzymes are under the control of the NF-E2 related factor 2 (Nrf2) transcription factor including members of the glutathione S-transferase (GST) and aldo-keto reductase (AKR) families.


METHODS
RT-PCR and Western immunoblotting were used to measure changes in transcript and protein levels in response to 5-h freezing exposure of hatchlings.


RESULTS
Transcript levels of Nrf2 increased in turtle brain, liver, and muscle by 1.5- to 2-fold, and protein levels increased in the brain and muscle by 1.6- to 2.3-fold in response to freezing. GSTs responded strongly to freezing in turtle brain with amounts of GSTP1, M1, K1, and A3 elevated by 1.5- to 2.4-fold. GSTM3 and T1 rose by 1.8- to 2.3-fold in gut, whereas reduced levels of GSTP1, M1, M3, and K1 were found in livers of frozen animals. Levels of the AKR1B4 isozyme rose 2.1-fold during freezing in brain.


CONCLUSIONS
Freezing triggered tissue-specific changes in the antioxidant defenses in C.pictamarginata organs.


CONCLUSIONS
These data indicate that activation of an antioxidant response is an important aspect of natural freeze tolerance in turtles.
Krivoruchko
Anastasia
A
Institute of Biochemistry and Department of Biology, Carleton University, Ottawa, Ontario, Canada K1S 5B6. krivoruchko@gmail.com
Storey
Kenneth B
KB
D000064:Acclimatization; D000595:Amino Acid Sequence; D000818:Animals; D000975:Antioxidants / Q000378:metabolism*; D015153:Blotting, Western; D002645:Chickens / Q000235:genetics; D017931:DNA Primers; D004531:Eggs; D005260:Female; D005615:Freezing; D005784:Gene Amplification; D006801:Humans; D008969:Molecular Sequence Data; D051267:NF-E2-Related Factor 2 / Q000737:chemistry / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D017386:Sequence Homology, Amino Acid; D014426:Turtles / Q000235:genetics / Q000502:physiology*; D014981:Xenopus / Q000235:genetics; D015027:Zebrafish / Q000235:genetics
D000975:Antioxidants; D017931:DNA Primers; D051267:NF-E2-Related Factor 2; D012313:RNA