Genetic compensation induced by deleterious mutations but not gene knockdowns

Zebrafish embryos injected with egfl7 morpholino exhibit severe vascular defects but egfl7 mutants do not show any obvious phenotypes, illustrating the power of comparing mutants and morphants to identify modifier genes. Mutant versus morphant phenotypes Antisense approaches to gene knockdown or interference, using agents such as siRNA and morpholino oligomers, have been criticized as being prone to off-target effects that can lead to phenotypes unrelated to the silencing of the target gene. Didier Stainer and colleagues contribute to this debate with a report that may cast doubt on the superiority of genetic inactivation versus knockdown. They show that 'morphant' zebrafish embryos in which the egfl7 gene is silenced using morpholinos, display severe vascular defects, whereas egfl7 mutant fish show very mild phenotypes. The discrepancy is a result of genetic compensation induced by deleterious mutations (upregulation of Emilins to counter the loss of Egfl7), but not by transcriptional or translational knockdown. This work illustrates the power of comparing mutants and morphants to identify modifier genes. Cells sense their environment and adapt to it by fine-tuning their transcriptome. Wired into this network of gene expression control are mechanisms to compensate for gene dosage. The increasing use of reverse genetics in zebrafish, and other model systems, has revealed profound differences between the phenotypes caused by genetic mutations and those caused by gene knockdowns at many loci 1 , 2 , 3 , an observation previously reported in mouse and Arabidopsis 4 , 5 , 6 , 7 . To identify the reasons underlying the phenotypic differences between mutants and knockdowns, we generated mutations in zebrafish egfl7 , an endothelial extracellular matrix gene of therapeutic interest, as well as in vegfaa . Here we show that egfl7 mutants do not show any obvious phenotypes while animals injected with egfl7 morpholino (morphants) exhibit severe vascular defects. We further observe that egfl7 mutants are less sensitive than their wild-type siblings to Egfl7 knockdown, arguing against residual protein function in the mutants or significant off-target effects of the morpholinos when used at a moderate dose. Comparing egfl7 mutant and morphant proteomes and transcriptomes, we identify a set of proteins and genes that are upregulated in mutants but not in morphants. Among them are extracellular matrix genes that can rescue egfl7 morphants, indicating that they could be c