Dataset Information

Knockdown of best1 Gene in Zebrafish Caused Abnormal Neuronal and Skeletal Development - A Subtype of Craniovertebral Junction Malformation?

ABSTRACT:

Objective

To investigate the developmental defects caused by knockdown of best1 gene in zebrafish as a model for a subtype of craniovertebral junction (CVJ) malformation.

Methods

Two antisense morpholinos (MOs) were designed targeting zebrafish best1 to block translation (ATG-MO) or to disrupt splicing (I3E4-MO). MOs were microinjected into fertilized one-cell embryos. Efficacy of splicing MO was confirmed by reverse transcription-polymerase chain reaction. Phenotypes were analyzed and quantified by microscopy at multiple developmental stages. Neuronal outgrowth was assessed in transgenic zebrafish expressing green fluorescent protein in neurons. Skeletal ossification was visualized by Calcein staining.

Results

Knockdown of best1 resulted in zebrafish embryos with shorter body length, curved axis, low survival rate, microcephaly, reduced eye size, smaller head and brain, impaired neuronal outgrowth, and reduced ossification of craniofacial and vertebral bone.

Conclusion

Best1 gene plays critical roles in ophthalmologic, neurological and skeletal development in zebrafish. A patient with a premature stop codon in BEST1 gene exhibited similar phenotypes, implying a subtype of CVJ malformation.

SUBMITTER: Liu Z

PROVIDER: S-EPMC11224734 | biostudies-literature | 2024 Jun

REPOSITORIES: biostudies-literature

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Publications

Knockdown of best1 Gene in Zebrafish Caused Abnormal Neuronal and Skeletal Development - A Subtype of Craniovertebral Junction Malformation?

Liu Zhenlei Z Li Kang K Wang Kai K Zhang Lei L Jia Shanhang S Wang He H Jian Fengzeng F Wu Hao H

Neurospine 20240201 2

<h4>Objective</h4>To investigate the developmental defects caused by knockdown of best1 gene in zebrafish as a model for a subtype of craniovertebral junction (CVJ) malformation.<h4>Methods</h4>Two antisense morpholinos (MOs) were designed targeting zebrafish best1 to block translation (ATG-MO) or to disrupt splicing (I3E4-MO). MOs were microinjected into fertilized one-cell embryos. Efficacy of splicing MO was confirmed by reverse transcription-polymerase chain reaction. Phenotypes were analyze ...[more]

PMID: 38317543

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Project description:Myosin-binding protein C (MyBPC) in the muscle sarcomere interacts with several contractile and structural proteins. Mutations in the cardiac isoform (MyBPC-3) in humans, or animal knockout, are associated with cardiomyopathy. Function of the fast skeletal isoform (MyBPC-2) in living muscles is less understood. This question was addressed using zebrafish models, combining gene expression data with functional analysis of contractility and small-angle x-ray diffraction measurements of filament structure. Fast skeletal MyBPC-2B, the major isoform, was knocked down by >50% using morpholino antisense nucleotides. These morphants exhibited a skeletal myopathy with elevated apoptosis and up-regulation of factors associated with muscle protein degradation. Morphant muscles had shorter sarcomeres with a broader length distribution, shorter actin filaments, and a wider interfilament spacing compared with controls, suggesting that fast skeletal MyBPC has a role in sarcomere assembly. Active force was reduced more than expected from the decrease in muscle size, suggesting that MyBPC-2 is required for optimal force generation at the cross-bridge level. The maximal shortening velocity was significantly increased in the MyBPC-2 morphants, but when related to the sarcomere length, the difference was smaller, reflecting that the decrease in MyBPC-2B content and the resulting myopathy were accompanied by only a minor influence on filament shortening kinetics. In the controls, equatorial patterns from small-angle x-ray scattering revealed that comparatively few cross-bridges are attached (as evaluated by the intensity ratio of the 11 and 10 equatorial reflections) during active contraction. X-ray scattering data from relaxed and contracting morphants were not significantly different from those in controls. However, the increase in the 11:10 intensity ratio in rigor was lower compared with that in controls, possibly reflecting effects of MyBPC on the cross-bridge interactions. In conclusion, lack of MyBPC-2 results in a severe skeletal myopathy with structural changes and muscle weakness.

Dataset Information

Knockdown of best1 Gene in Zebrafish Caused Abnormal Neuronal and Skeletal Development - A Subtype of Craniovertebral Junction Malformation?

Objective

Methods

Results

Conclusion

Publications

Knockdown of best1 Gene in Zebrafish Caused Abnormal Neuronal and Skeletal Development - A Subtype of Craniovertebral Junction Malformation?

Similar Datasets

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