A dual‐color marker system for in vivo visualization of cell cycle progression in A rabidopsis

• Published in: The Plant journal : for cell and molecular biology 2014-11, Vol.80 (3), p.541-552
• Main Authors: Yin, Ke, Ueda, Minako, Takagi, Hitomi, Kajihara, Takehiro, Sugamata Aki, Shiori, Nobusawa, Takashi, Umeda‐Hara, Chikage, Umeda, Masaaki
• Format: Article
• Language: English
• ISSN: 0960-7412; 1365-313X
• DOI: 10.1111/tpj.12652

Visualization of the spatiotemporal pattern of cell division is crucial to understand how multicellular organisms develop and how they modify their growth in response to varying environmental conditions. The mitotic cell cycle consists of four phases: S ( DNA replication), M (mitosis and cytokinesis), and the intervening G 1 and G 2 phases; however, only G 2/ M ‐specific markers are currently available in plants, making it difficult to measure cell cycle duration and to analyze changes in cell cycle progression in living tissues. Here, we developed another cell cycle marker that labels S ‐phase cells by manipulating A rabidopsis CDT 1a , which functions in DNA replication origin licensing. Truncations of the CDT 1a coding sequence revealed that its carboxy‐terminal region is responsible for proteasome‐mediated degradation at late G 2 or in early mitosis. We therefore expressed this region as a red fluorescent protein fusion protein under the S ‐specific promoter of a histone 3.1‐type gene, HISTONE THREE RELATED 2 ( HTR 2 ), to generate an S / G 2 marker. Combining this marker with the G 2/ M ‐specific CYCB 1 ‐ GFP marker enabled us to visualize both S to G 2 and G 2 to M cell cycle stages, and thus yielded an essential tool for time‐lapse imaging of cell cycle progression. The resultant dual‐color marker system, Cell Cycle Tracking in Plant Cells ( C ytrap), also allowed us to identify root cells in the last mitotic cell cycle before they entered the endocycle. Our results demonstrate that C ytrap is a powerful tool for in vivo monitoring of the plant cell cycle, and thus for deepening our understanding of cell cycle regulation in particular cell types during organ development.