asb5a/asb5b Double Knockout Affects Zebrafish Cardiac Contractile Function.

Area of Science:

• Developmental Biology
• Genetics
• Cardiovascular Research

Background:

• Ankyrin repeat and suppression-of-cytokine-signaling box (Asb) proteins are ubiquitin ligase E3s.
• Asb5 has six ankyrin-repeat domains, and zebrafish possess two gene isoforms: asb5a and asb5b.
• The roles of asb5 gene inactivation in zebrafish development and heart function remain uncharacterized.

Purpose of the Study:

• To investigate the effects of asb5 gene inactivation on zebrafish embryonic development and heart function.
• To explore the functional redundancy between asb5a and asb5b isoforms.
• To elucidate the molecular mechanisms underlying asb5's role in cardiac development.

Main Methods:

• CRISPR/Cas9 was used to generate asb5a-knockout zebrafish.
• Morpholino interference targeted asb5b in wild-type and asb5a-knockout zebrafish.
• RNA-sequencing, cluster analysis, heartbeat analysis, and bioinformatics tools (STRING, Cytoscape) were employed.

Main Results:

• Single asb5a knockout showed no abnormal phenotypes; asb5b knockdown in wild-type also had no impact.
• Simultaneous asb5a knockout and asb5b knockdown resulted in severe pericardial cavity enlargement and atrial dilation.
• RNA-seq identified enriched cardiac muscle contraction genes, and heartbeat analysis revealed significant functional changes in double knockouts.
• Bioinformatic analyses pinpointed 11 cardiac-contraction-related hub genes and potential regulation via calcium ion channels.

Conclusions:

• Zebrafish asb5a and asb5b exhibit functional redundancy.
• The simultaneous inactivation of both asb5 isoforms significantly impairs early zebrafish heart development and contractile function.
• This study provides critical insights into the mechanism of asb5 in cardiac regulation.