Dissection of zebrafish shha function using site-specific targeting with a Cre-dependent genetic switch

ELife
Kotaro SugimotoKazu Kikuchi

Abstract

Despite the extensive use of zebrafish as a model organism in developmental biology and regeneration research, genetic techniques enabling conditional analysis of gene function are limited. In this study, we generated Zwitch, a Cre-dependent invertible gene-trap cassette, enabling the establishment of conditional alleles in zebrafish by generating intronic insertions via in vivo homologous recombination. To demonstrate the utility of Zwitch, we generated a conditional sonic hedgehog a (shha) allele. Homozygous shha mutants developed normally; however, shha mutant embryos globally expressing Cre exhibited strong reductions in endogenous shha and shha target gene mRNA levels and developmental defects associated with null shha mutations. Analyzing a conditional shha mutant generated using an epicardium-specific inducible Cre driver revealed unique roles for epicardium-derived Shha in myocardial proliferation during heart development and regeneration. Zwitch will extend the utility of zebrafish in organ development and regeneration research and might be applicable to other model organisms.

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Citations

Apr 14, 2019·Developmental Dynamics : an Official Publication of the American Association of Anatomists·Stone ElworthyTimothy J A Chico
Jun 29, 2018·Nature Reviews. Cardiology·Jingli Cao, Kenneth D Poss
Feb 14, 2021·Journal of Cardiovascular Development and Disease·Cassie L KemmlerChristian Mosimann
Apr 10, 2021·Science·Masahito OgawaKazu Kikuchi

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Datasets Mentioned

BETA
30269

Methods Mentioned

BETA
gene knockout
gene trap
PCR
Genotyping
fluorescence-activated
transgenic
FACS
amputation
electrophoresis
cleavage assay

Software Mentioned

TAL Effector Nucleotide Targeter
Zwitch
Ensembl
ImageJ
pZwitch

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