|ZFIN ID: ZDB-PUB-091120-71|
Illuminating cell-cycle progression in the developing zebrafish embryo
Sugiyama, M., Sakaue-Sawano, A., Iimura, T., Fukami, K., Kitaguchi, T., Kawakami, K., Okamoto, H., Higashijima, S.I., and Miyawaki, A.
|Source:||Proceedings of the National Academy of Sciences of the United States of America 106(49): 20812-20817 (Journal)|
|Registered Authors:||Higashijima, Shin-ichi, Kawakami, Koichi, Miyawaki, Atsushi, Okamoto, Hitoshi, Sugiyama, Mayu|
|Keywords:||cell cycle, fluorescent protein, imaging, ubiquitination|
|PubMed:||19923430 Full text @ Proc. Natl. Acad. Sci. USA|
Sugiyama, M., Sakaue-Sawano, A., Iimura, T., Fukami, K., Kitaguchi, T., Kawakami, K., Okamoto, H., Higashijima, S.I., and Miyawaki, A. (2009) Illuminating cell-cycle progression in the developing zebrafish embryo. Proceedings of the National Academy of Sciences of the United States of America. 106(49):20812-20817.
ABSTRACTBy 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.