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Fig. 1 Identification of radiosensitizing mutation #7.
(A) Wild-type AB strain embryos were exposed to the indicated amount of IR at 24 hpf and visualized by brightfield microscopy 6 hours later. A threshold of IR exists between 8 and 15 Gy that gives rise to obvious cell death (seen as opaque tissue, marked by arrowheads) in the brain. (B) Wild-type embryos (derived from crossing wild-type parents) or rs7 mutant embryos (derived from crossing rs7 heterozygous parents) were exposed to 8 Gy IR at 24 hpf and visualized by brightfield microscopy 6 hours later. Arrowheads in the rs7 mutant mark cell death reminiscent of exposure of wild-type embryos to 15 Gy (as shown in A) (C) Embryos in (B) were sorted by phenotype, fixed at 6 hpIR and analyzed by immunofluorescence to detect activated Caspase-3. The “curly-up” tail phenotype and opaque tissue in the head were used to identify rs7 mutants. These phenotypes are present (to different degrees of severity) in both unirradiated and irradiated mutants such that they can be readily distinguished from siblings and wild-type at 30 hpf. After fixation of mutants, tails were clipped to distinguish them from wild-type embryos, which were analyzed in the same tube for Caspase-3 activity. Embryos were grouped for analysis according to whether embryos were irradiated or not, and rs7 mutants were identified based on the presence of tail-clips. Arrowheads point to the enhanced apoptosis in the spinal cord of an rs7 mutant, and arrows point to increased apoptosis in the ICM region of an rs7 mutant. (D) Immunofluorescence in the spinal cords from at least 10 embryos per group in (C) was quantified. wt; wild-type. See also.