Fig. 3

Zebrafish Chop Is Dispensable for Rapid ER Stress-Induced Apoptosis, but Is Required for 24 Hr Apoptosis in the Caudal Fin Fold

(A) Alignment of zebrafish Chop with human and mouse orthologs. Consensus sequence is listed below the aligned sequences.

(B–D) DMSO-treated embryos have minimal chop expression, while embryos treated with thapsigargin for 3 hr (C) have high levels of chop expression in the tail (brackets), including the tail epidermis (black arrowhead). Expression was also observed in the dorsal trunk epidermis (open arrowhead in C).

(E) An anterior view reveals additional chop expression in the epiphysis region (dashed white box) and the lens (black arrowhead). Images in (B)–(E) represent 100% of embryos examined; n e 14 embryos per condition.

(F) Agarose gel (1%) showing alternate splice product generated 28 hr after chop morpholino (MO) injection compared to uninjected controls (Ctrl.); both groups of embryos were treated with thapsigargin at 24 hpf.

(G) Depiction of splice-blocking event in chop morphant mRNA compared to wild-type, as determined by sequencing.

(H) chop morpholino (5 ng) was injected into p53 homozygous mutant embryos, which were then grown to 24 hpf, treated with thapsigargin for 4 hr, and assayed for cell death by AO staining. Cropped photos were quantitated for total fluorescence intensity with Volocity software, and values were normalized to controls.

(I) chop morpholino-injected embryos were grown to 24 hpf, treated with thapsigargin for 4 hr, washed into egg medium, left for 24 hr, and then stained with AO and assayed as in (H). All data are represented as means ± SEM.