Zebrafish atoh8 mutants do not recapitulate morpholino phenotypes

Place, Elsie S.; Smith, James C.

doi:10.1371/journal.pone.0171143

Cited by 16 publications

(20 citation statements)

References 39 publications

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“…We have identified compensation triggered by a mutation in actc1b but not following morpholino mediated knockdown of Actc1b. There has been considerable debate recently as a result of phenotypic differences between mutant lines and morpholino-mediated knockdown [ 19 – 22 ]. The study by Rossi et al (2015) provided an example where this difference in phenotype was due to compensation, rather than morpholino off target effects as previously suggested.…”

Section: Discussionmentioning

confidence: 99%

Genetic compensation triggered by actin mutation prevents the muscle damage caused by loss of actin protein

et al. 2018

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The lack of a mutant phenotype in homozygous mutant individuals’ due to compensatory gene expression triggered upstream of protein function has been identified as genetic compensation. Whilst this intriguing process has been recognized in zebrafish, the presence of homozygous loss of function mutations in healthy human individuals suggests that compensation may not be restricted to this model. Loss of skeletal α-actin results in nemaline myopathy and we have previously shown that the pathological symptoms of the disease and reduction in muscle performance are recapitulated in a zebrafish antisense morpholino knockdown model. Here we reveal that a genetic actc1b mutant exhibits mild muscle defects and is unaffected by injection of the actc1b targeting morpholino. We further show that the milder phenotype results from a compensatory transcriptional upregulation of an actin paralogue providing a novel approach to be explored for the treatment of actin myopathy. Our findings provide further evidence that genetic compensation may influence the penetrance of disease-causing mutations.

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Section: Discussionmentioning

confidence: 99%

Genetic compensation triggered by actin mutation prevents the muscle damage caused by loss of actin protein

et al. 2018

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“…Discrepant mutant and morphant phenotypes have been reported for an ever-increasing number of genes, raising several caveats about the use of MOs [ 49 ] [ 50 ] [ 51 ] [ 52 ] [ 53 ] [ 54 ]. Explanations to account for MOs not replicating null-mutant phenotypes include off-target/non-specific effects of MOs, unexpected changes in gene expression, and apoptosis via a p53-dependent mechanism [ 55 ] [ 49 ] [ 50 ] [ 51 ] [ 56 ].…”

Section: Discussionmentioning

confidence: 99%

Investigation of Islet2a function in zebrafish embryos: Mutants and morphants differ in morphologic phenotypes and gene expression

et al. 2018

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Zebrafish primary motor neurons differ from each other with respect to morphology, muscle targets and electrophysiological properties. For example, CaP has 2-3-fold larger densities of both inward and outward currents than do other motor neurons. We tested whether the transcription factor Islet2a, uniquely expressed in CaP, but not other primary motor neurons, plays a role in specifying its stereotypic electrophysiological properties. We used both TALEN-based gene editing and antisense morpholino approaches to disrupt Islet2a function. Our electrophysiology results do not support a specific role for Islet2a in determining CaP’s unique electrical properties. However, we also found that the morphological phenotypes of CaP and a later-born motor neuron differed between islet2a mutants and morphants. Using microarrays, we tested whether the gene expression profiles of whole embryo morphants, mutants and controls also differed. Morphants had 174 and 201 genes that were differentially expressed compared to mutants and controls, respectively. Further, islet2a was identified as a differentially expressed gene. To examine how mutation of islet2a affected islet gene expression specifically in CaPs, we performed RNA in situ hybridization. We detected no obvious differences in expression of islet1, islet2a, or islet2b in CaPs of mutant versus sibling control embryos. However, immunolabeling studies revealed that an Islet protein persisted in CaPs of mutants, albeit at a reduced level compared to controls. While we cannot exclude requirement for some Islet protein, we conclude that differentiation of the CaP’s stereotypic large inward and outward currents does not have a specific requirement for Islet2a.

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“…Two types of cldn5a morphants showed similar phenotypes of disrupted heart laterality and exogenous RNA rescued the cldn5a morpholino-induced phenotypes. Several recent papers reported that genome-engineered mutants failed to recapitulate the morpholino-induced phenotypes [ 54 – 56 ]. Although genetic compensation might be induced in mutant embryos [ 57 ], morpholino-based studies should be carefully considered whether their phenotypes are driven by off-target effects.…”

Section: Discussionmentioning

confidence: 99%

Claudin5a is required for proper inflation of Kupffer's vesicle lumen and organ laterality

et al. 2017

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Left-right asymmetric organ development is critical to establish a proper body plan of vertebrates. In zebrafish, the Kupffer’s vesicle (KV) is a fluid-filled sac which controls asymmetric organ development, and a properly inflated KV lumen by means of fluid influx is a prerequisite for the asymmetric signal transmission. However, little is known about the components that support the paracellular tightness between the KV luminal epithelial cells to sustain hydrostatic pressure during KV lumen expansion. Here, we identified that the claudin5a (cldn5a) is highly expressed at the apical surface of KV epithelial cells and tightly seals the KV lumen. Downregulation of cldn5a in zebrafish showed a failure in organ laterality that resulted from malformed KV. In addition, accelerated fluid influx into KV by combined treatment of forskolin and 3-isobutyl-1-methylxanthine failed to expand the partially-formed KV lumen in cldn5a morphants. However, malformed KV lumen and defective heart laterality in cldn5a morphants were significantly rescued by exogenous cldn5a mRNA, suggesting that the tightness between the luminal epithelial cells is important for KV lumen formation. Taken together, these findings suggest that cldn5a is required for KV lumen inflation and left-right asymmetric organ development.

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Zebrafish atoh8 mutants do not recapitulate morpholino phenotypes

Cited by 16 publications

References 39 publications

Genetic compensation triggered by actin mutation prevents the muscle damage caused by loss of actin protein

Genetic compensation triggered by actin mutation prevents the muscle damage caused by loss of actin protein

Investigation of Islet2a function in zebrafish embryos: Mutants and morphants differ in morphologic phenotypes and gene expression

Claudin5a is required for proper inflation of Kupffer's vesicle lumen and organ laterality

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