Lineage labeling with zebrafish hand2 Cre and CreERT2 recombinase CRISPR knock-ins

Area of Science:

• Developmental Biology
• Genetics
• Zebrafish Models

Background:

• CRISPR-Cas9 knock-in technology facilitates endogenous Cre recombinase driver generation.
• This enables lineage tracing and cell type-specific gene studies.
• Endogenous zebrafish hand2 Cre and CreERT2 drivers were generated using GeneWeld CRISPR-Cas9 precision targeted integration.

Purpose of the Study:

• To generate and characterize endogenous zebrafish hand2 Cre and CreERT2 drivers.
• To utilize these drivers for lineage tracing during embryogenesis and larval organogenesis.
• To investigate the contribution of hand2-expressing progenitors to various tissues.

Main Methods:

• CRISPR-Cas9 knock-in technology (GeneWeld) for generating endogenous drivers.
• Crossed hand2-2A-cre and hand2-2A-creERT2 knock-ins with loxP-based Switch reporters.
• Temporal induction of CreERT2 activity at specific embryonic and larval stages.

Main Results:

• Broad labeling of mesodermal and neural crest-derived lineages, including branchial arches, heart, liver, intestine, and enteric neurons.
• Novel hand2 lineage tracing patterns observed in venous blood vessels.
• Temporal analysis revealed hand2 progenitor contributions to venous and intestinal vasculature (24-48hpf) and restricted larval lineages.

Conclusions:

• hand2 progenitors from lateral plate mesoderm and ectoderm contribute to diverse embryonic lineages.
• hand2-expressing cells show restricted lineage contributions in later larval stages.
• The developed endogenous hand2 Cre and CreERT2 drivers are valuable tools for studying hand2 lineages in zebrafish development.