PUBLICATION

Control of cardiac contractions using Cre-lox and degron strategies in zebrafish

Authors
Juan, T., Bellec, M., Cardoso, B., Athéa, H., Fukuda, N., Albu, M., Günther, S., Looso, M., Stainier, D.Y.R.
ID
ZDB-PUB-240110-16
Date
2024
Source
Proceedings of the National Academy of Sciences of the United States of America   121: e2309842121e2309842121 (Journal)
Registered Authors
Albu, Marga, Juan, Thomas, Stainier, Didier
Keywords
Cre-lox, Degron, cardiac contractions, cardiac troponin T, cpFRB2-FKBP
Datasets
GEO:GSE232835
MeSH Terms
  • Alleles
  • Animals
  • Perciformes*
  • Degrons
  • Myocytes, Cardiac
  • Zebrafish*/genetics
(all 6)
PubMed
38194447 Full text @ Proc. Natl. Acad. Sci. USA
Abstract
Cardiac contractions and hemodynamic forces are essential for organ development and homeostasis. Control over cardiac contractions can be achieved pharmacologically or optogenetically. However, these approaches lack specificity or require direct access to the heart. Here, we compare two genetic approaches to control cardiac contractions by modulating the levels of the essential sarcomeric protein Tnnt2a in zebrafish. We first recombine a newly generated tnnt2a floxed allele using multiple lines expressing Cre under the control of cardiomyocyte-specific promoters, and show that it does not recapitulate the tnnt2a/silent heart mutant phenotype in embryos. We show that this lack of early cardiac contraction defects is due, at least in part, to the long half-life of tnnt2a mRNA, which masks the gene deletion effects until the early larval stages. We then generate an endogenous Tnnt2a-eGFP fusion line that we use together with the zGRAD system to efficiently degrade Tnnt2a in all cardiomyocytes. Using single-cell transcriptomics, we find that Tnnt2a depletion leads to cardiac phenotypes similar to those observed in tnnt2a mutants, with a loss of blood and pericardial flow-dependent cell types. Furthermore, we achieve conditional degradation of Tnnt2a-eGFP by splitting the zGRAD protein into two fragments that, when combined with the cpFRB2-FKBP system, can be reassembled upon rapamycin treatment. Thus, this Tnnt2a degradation line enables non-invasive control of cardiac contractions with high spatial and temporal specificity and will help further understand how they shape organ development and homeostasis.
Genes / Markers
Figures
Figure Gallery (6 images)
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Expression
No data available
Phenotype
No data available
Mutations / Transgenics
Allele Construct Type Affected Genomic Region
bns141TgTransgenic Insertion
    bns511PtTransgenic Insertion
    bns513PtTransgenic Insertion
    bns594TgTransgenic Insertion
      bns625TgTransgenic Insertion
      bns626TgTransgenic Insertion
        bns672TgTransgenic Insertion
          bns704TgTransgenic Insertion
            hsc185TgTransgenic Insertion
              mn0031GtTransgenic Insertion
              1 - 10 of 16
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              Human Disease / Model
              No data available
              Sequence Targeting Reagents
              Target Reagent Reagent Type
              tnnt2aCRISPR12-tnnt2aCRISPR
              tnnt2aCRISPR13-tnnt2aCRISPR
              1 - 2 of 2
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              Fish
              No data available
              Antibodies
              Name Type Antigen Genes Isotypes Host Organism
              Ab1-actbmonoclonal
                		IgG1Mouse
                1 - 1 of 1
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                Orthology
                No data available
                Engineered Foreign Genes
                Marker Marker Type Name
                cpFRB2-FKBPEFGcpFRB2-FKBP
                CreEFGCre
                DsRedEFGDsRed
                EGFPEFGEGFP
                mCherryEFGmCherry
                mRFP1EFGmRFP1
                TagBFPEFGTagBFP
                TomatoEFGTomato
                zGRADEFGzGRAD
                1 - 9 of 9
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                Mapping
                No data available
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                Citation
                Juan, T., Bellec, M., Cardoso, B., Athéa, H., Fukuda, N., Albu, M., Günther, S., Looso, M., Stainier, D.Y.R. (2024) Control of cardiac contractions using Cre-lox and degron strategies in zebrafish. Proceedings of the National Academy of Sciences of the United States of America. 121:e2309842121e2309842121.