Fig. 1

Expression of exogenous GFP and endogenous chop transcripts and the appearance of HrRCs in the brain of hypoxia-exposed huORFZ embryos. (A) Schematic illustration of experimental workflow. Zebrafish embryos from transgenic line huORFZ developed at 72 hpf, 2 h prior to oxygen recovery (R-2), were exposed to hypoxia for 2 h, followed by the start of oxygen recovery (R0, 74 hpf). (B) No GFP signal was presented in the huORFZ embryos served as mock control groups at 72 hpf and R0 (74 hpf). (C) The GFP signal started to appear in embryos undergoing oxygen recovery for 12 h (R12), while GFP was observed in the brain at R15. (D) Dorsal and lateral views of the temporospatial expression of chop transcript in the nonhypoxia-exposure huORFZ (normoxia; positive control group) embryos at 98 hpf (equivalent to R24) and hypoxia-exposure huORFZ embryos at R24 (equivalent to 98 hpf) and R36 (equivalent to 110 hpf) using whole-mount in situ hybridization (WISH). The chop mRNA was mainly expressed in the telencephalon (Tel), habenula (Hb), optic tectum (TeO), ventricular zone (VZ), and medulla oblongata (MO). Western blot analyses of (E) CHOP and Endouc proteins presented at various stages of normoxia- and hypoxia-exposure huORFZ embryos. Total proteins were extracted from the heads of embryos at the start of recovery (74 hpf; R0) through 146 hpf (R72), as indicated. The control group consisted of untreated huORFZ embryos (normoxia) after extraction of total proteins from the head at the corresponding hpf of treated embryos. α-Tubulin served as a loading control. HrRCs: hypoxia-responsive recovering cells; hpf: hours postfertilization.