Tracing the Silhouette of Individual Cells in S/G2/M Phases with Fluorescence

Abstract

The APC(Cdh1) E3 ligase is active in the late M and G(1) phases. Geminin is a direct substrate of the APC(Cdh1) complex, and accumulates during the S, G(2), and M phases. By fusing the amino-terminal region of Geminin to fluorescent proteins, we have developed cell cycle markers that accumulate in the S/G(2)/M phases in both the nucleus and the cytoplasm. These markers reveal the morphology of individual cells that have undergone DNA replication, allowing us to monitor cell growth relative to differentiation of various cell types. After electroporating the developing mouse embryos, we highlighted neuroepithelial progenitors in the S/G(2)/M phases, which possessed an elongated morphology with an apical and/or a basal attachment. We also show that nuclear localization of the ubiquitin ligase for Geminin is essential for full performance of the markers.