Fig. 5

Effects of HEP induction on early aspects of zebrafish development. Embryos from crosses of wild-type and HEP^+/- transgenic (Tg) fish were exposed to high temperature for 1 h and then allowed to develop at a lower temperature to appropriate stages. (A, B) Induction of HEP at the high/oblong stage led to epiboly arrest and thickening of the blastoderm in about half of the embryos (B) when the remaining embryos were normal and reached 80% epiboly (A). (C–H) Induction of HEP at the sphere stage led to elongation of embryos along the animal–vegetal axis at the bud stage in about half of the embryos (D, F), while the remaining embryos were normal (C, E). By 26 hpf, many of elongated embryos had died and live embryos were severely disrupted in their posterior region (compare G and H). Scale bar, 200 µm. (I, J) Genotyping of embryos exposed to heat shock at the sphere stage. G, genomic sequences amplified for positive control; H, HEP-derived sequences showing that the embryos harbored the HEP gene; M, size markers (500 and 200 bp). (I) Embryos that were normal or abnormal in terms of morphology and DV marker expression were subjected to genotyping. The data demonstrated that normal embryos were negative but abnormal embryos were positive for HEP. (J) Offspring embryos from crosses of HEP^+/- Tg and wild-type fish were injected with pou2 mRNA and then given heat shock at the sphere stage. Genotyping showed that the embryos with expanded eve1 expression were negative (+++; Table 2), and the embryos with slightly reduced expression (+) were positive for HEP.