Bakunaite, E. | Search bioRxiv

bioRxiv2025

Epicardial Tcf21 facilitates cardiomyocyte dedifferentiation and heart regeneration in zebrafish

Unlike mammals, zebrafish (Danio rerio) are able to regenerate their hearts after injury, making them an excellent model organism for studying the molecular mechanisms underlying heart regeneration. Epicardium, the outermost layer of the heart, is an essential player in this process. Injury-induced epicardium activation, characterized by the expression of embryonic epicardial marker genes including tcf21 supports cardiac regeneration by providing various cell types and releasing paracrine signals that promote the restoration of damaged tissue. However, the molecular mechanisms involved in this process are insufficiently understood. In this study, we describe a conditional tcf21flox allele and use it to investigate the role of Tcf21 in heart regeneration. By employing 4-hydroxytamoxifen inducible CreERT2 recombinase, we eliminated tcf21 expression in adult fish. Our findings indicate that loss of this transcription factor reduces the presence of dedifferentiated cardiomyocytes in the injury area and impairs heart regeneration. This work provides new insights into the molecular basis of the epicardial response to heart injury and its role in guiding heart regeneration.

developmental biology↗

bioRxiv2024

Highly efficient tamoxifen-inducible Cre recombination in embryonic, larval and adult zebrafish

We have generated transgenic lines containing zebrafish-optimized CreERT2 recombinase under the control of a recombinant ubbR promoter consisting of the zebrafish ubiquitin promoter supplemented with an intronic enhancer from the carp beta-actin2 gene. These lines enable highly efficient tamoxifen-inducible recombination in embryonic, larval and adult zebrafish. AbstractThe ability to inactivate gene function in an adult organism is essential for studies of biological processes such as regeneration and behavior. This is best achieved by engineering an allele which could be conditionally inactivated using Cre recombinase and subsequently inactivating gene function using a drug-inducible Cre recombinase. Several recent studies clearly demonstrate feasibility of engineering such conditional alleles in zebrafish. Meanwhile, achieving sufficient degree of recombination to induce complete loss of function has remained a major limitation. Herein we address this limitation by engineering a recombinant ubiquitin promoter ubbR consisting of the zebrafish ubiquitin promoter supplemented with an intronic enhancer from the carp beta-actin2 gene. Using phiC31-mediated targeted integration, we demonstrate that ubbR clearly outperforms both parental promoters as well as currently available ubiquitous CreERT2 driver lines at all embryonic and larval stages tested. Furthermore, the ubbR:CreERT2driver line we generated enables near-complete inactivation of floxed alleles in adult zebrafish hearts. Finally, we demonstrate that our ubbRpromoter retains high activity when integrated at other genomic loci, making it uniquely suitable for robust expression of transgenes at all stages of zebrafish ontogenesis. HighlightsO_LIUsed targeted integration to directly compare different CreERT2 drivers C_LIO_LIGenerated a ubiquitous ubbR:CreERT2 driver line capable of near-complete inactivation of floxed genes in adult zebrafish hearts C_LIO_LIDemonstrated that the recombinant ubbR promoter is suitable for robust transgene expression when integrated at different genomic loci C_LI

developmental biology↗