Related Experiment Video
Updated: Jun 18, 2026
Light Sheet Microscopy Imaging and Mounting Strategies for Early Zebrafish Embryos
Published on: July 19, 2024
Illuminating cell-cycle progression in the developing zebrafish embryo
Mayu Sugiyama1, Asako Sakaue-Sawano, Tadahiro Iimura
1Laboratory for Cell Function and Dynamics, Advanced Technology Development Group, Brain Science Institute, RIKEN, 2-1 Hirosawa, Wako-city, Saitama 351-0198, Japan.
Abstract:
By exploiting the cell-cycle-dependent proteolysis of two ubiquitination oscillators, human Cdt1 and geminin, which are the direct substrates of SCF(Skp2) and APC(Cdh1) complexes, respectively, Fucci technique labels mammalian cell nuclei in G(1) and S/G(2)/M phases with different colors. Transgenic mice expressing these G(1) and S/G(2)/M markers offer a powerful means to investigate the coordination of the cell cycle with morphogenetic processes. We attempted to introduce these markers into zebrafish embryos to take advantage of their favorable optical properties. However, although the fundamental mechanisms for cell-cycle control appear to be well conserved among species, the G(1) marker based on the SCF(Skp2)-mediated degradation of human Cdt1 did not work in fish cells, probably because the marker was not ubiquitinated properly by a fish E3 ligase complex. We describe here the generation of a Fucci derivative using zebrafish homologs of Cdt1 and geminin, which provides sweeping views of cell proliferation in whole fish embryos. Remarkably, we discovered two anterior-to-posterior waves of cell-cycle transitions, G(1)/S and M/G(1), in the differentiating notochord. Our study demonstrates the effectiveness of using the Cul4(Ddb1)-mediated Cdt1 degradation pathway common to all metazoans for the development of a G(1) marker that works in the nonmammalian animal model.
Insights
Researchers developed a new Fucci system for zebrafish, enabling visualization of cell cycle progression. This system uses zebrafish proteins to track cell division dynamics in developing embryos, revealing novel cell-cycle transition waves.
Area of Science:
- Developmental Biology
- Cell Biology
- Molecular Biology
Background:
- The Fucci technique visualizes cell cycle phases (G1, S/G2/M) using fluorescent markers based on Cdt1 and geminin.
- Transgenic mice expressing Fucci markers aid in studying cell cycle coordination with morphogenesis.
- Adapting Fucci for zebrafish offers optical advantages but faced challenges with mammalian markers.
Purpose of the Study:
- To develop a functional Fucci derivative for zebrafish embryos using homologous proteins.
- To visualize cell proliferation and cell-cycle transitions in developing zebrafish.
- To investigate the conservation and adaptation of cell-cycle regulatory pathways across species.
Main Methods:
- Generated a novel Fucci system using zebrafish homologs of Cdt1 and geminin.
- Utilized the Cul4(Ddb1)-mediated Cdt1 degradation pathway for G1 marker development.
- Observed cell proliferation dynamics in whole zebrafish embryos.
Main Results:
- Successfully created a zebrafish-specific Fucci system for live imaging of cell proliferation.
- Identified two distinct anterior-to-posterior waves of cell-cycle transitions (G1/S and M/G1) in the notochord.
- Demonstrated the efficacy of a conserved Cdt1 degradation pathway for non-mammalian Fucci development.
Conclusions:
- A novel Fucci system adapted for zebrafish enables comprehensive visualization of cell cycle dynamics.
- The study reveals previously uncharacterized spatiotemporal patterns of cell-cycle progression during zebrafish development.
- Conserved metazoan Cdt1 degradation pathways are effective for engineering cross-species cell cycle reporters.
