Morpholino artifacts provide pitfalls and reveal a novel role for pro-apoptotic genes in hindbrain boundary development

► Previous studies have shown that antisense morpholino oligonucleotides (MOs) can have off-target effects mediated by activation of Tp53 and associated apoptosis. ► We show that in the zebrafish hindbrain, MOs can also non-specifically induce ectopic boundary marker gene expression. ► The induction of ectopic gene expression by MOs is due to a normal non-apoptotic role of the Bcl-caspase pathway that is required for hindbrain boundary gene expression. ► These findings reveal that a previous study suggesting that Wnt1 promotes neurogenesis and restricts hindbrain boundary marker expression is incorrect. ► The finding that MOs can induce non-apoptotic roles of the pro-apoptotic pathway further emphasizes the need for precautions in their use and interpretation of results.