Acute hypoxia effects on Keap1/Nrf2 (Mafs)-GST pathway related oxidative metabolism in muscle of Japanese flounder (Paralichthys olivaceus)

Acute hypoxia can aggravate the oxidation metabolism of fish muscle tissue. However, the molecular mechanism of oxidative metabolism in fish muscle under acute hypoxia is not very clear. We carried out effects of a typical oxidative metabolism pathway Keap1/Nrf2 (MafG)-GST on muscle oxidative metabolism of Japanese flounder (Paralichthys olivaceus) during acute hypoxia stimulation (1.65 ± 0.05 mg/L; 1 h, 3 h, 6 h, 12 h, 24 h) and reoxygenation (7.30 ± 0.08 mg/L; R12 h, R24 h, R48 h). The mRNAs of Nrf2 and GST in skeletal muscle were found co-existent, and their expressions were significant increase in 3 h and 6 h. The methylation level of CpG island1 in Nrf2 promoter, whose minimum value appeared at 3 h hypoxia treatment group, was affected by acute hypoxia, and it was negatively correlated with Nrf2 expression. The result suggests that environmental factors may regulate gene expression by epigenetic modification. Dual-luciferase reporter assay showed that GST gene was activated by transcription factor Nrf2, whose transcriptional activation binding region in GST promoter was antioxidant response element located near −980 and −852 sites, and Keap1 and MafG were Nrf2 antagonistic and synergistic factor, respectively. Furthermore, the GST activity changed with hypoxia and reoxygenation treatment in muscle, where other oxidative stress factor (MDA), antioxidant factors (T-AOC, GSH) and antioxidant enzyme activities (GST, SOD, CAT) were also changed. The results of MDA and T-AOC being further different between its hypoxia and normoxia groups (P < 0.05) at 6 h demonstrated that hypoxia stimulation lasting for 6 h would deeply affect Japanese flounder. The study illustrated that Japanese flounder responded to acute hypoxia in multiple metabolic levels by changing methylation status and transcription factor activation. It is significant to understand oxidative metabolic mechanism, analyze organism stress response and promote the scientific development of aquaculture.