Quantitative Cell Cycle Analysis Based on an Endogenous All-in-One Reporter for Cell Tracking and Classification

Publication date: 30 May 2017
Source:Cell Reports, Volume 19, Issue 9
Author(s): Thomas Zerjatke, Igor A. Gak, Dilyana Kirova, Markus Fuhrmann, Katrin Daniel, Magdalena Gonciarz, Doris Müller, Ingmar Glauche, Jörg Mansfeld
Cell cycle kinetics are crucial to cell fate decisions. Although live imaging has provided extensive insights into this relationship at the single-cell level, the limited number of fluorescent markers that can be used in a single experiment has hindered efforts to link the dynamics of individual proteins responsible for decision making directly to cell cycle progression. Here, we present fluorescently tagged endogenous proliferating cell nuclear antigen (PCNA) as an all-in-one cell cycle reporter that allows simultaneous analysis of cell cycle progression, including the transition into quiescence, and the dynamics of individual fate determinants. We also provide an image analysis pipeline for automated segmentation, tracking, and classification of all cell cycle phases. Combining the all-in-one reporter with labeled endogenous cyclin D1 and p21 as prime examples of cell-cycle-regulated fate determinants, we show how cell cycle and quantitative protein dynamics can be simultaneously extracted to gain insights into G1 phase regulation and responses to perturbations.

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Teaser

Zerjatke et al. present endogenously tagged PCNA as an all-in-one cell cycle reporter for living cells to classify all cell cycle phases and quiescence using a single fluorescent channel. Visualizing endogenous cyclin D1 in living cells, they show that cyclin D1 maintains G1 phase and prevents the transition into quiescence.


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