Abstract
Development, tissue homeostasis and tumor suppression depend critically on the correct regulation of cell division. Central in the cell division process is the decision whether to enter the next cell cycle and commit to going through the S and M phases, or to remain temporarily or permanently arrested. Cell cycle studies in genetic model systems could greatly benefit from visualizing cell cycle commitment in individual cells without the need of fixation. Here, we report the development and characterization of a reporter to monitor cell cycle entry in the nematode C. elegans. This reporter combines the mcm-4 promoter, to reveal Rb/E2F-mediated transcriptional control, and a live-cell sensor for CDK-activity. The CDK sensor was recently developed for use in human cells and consists of a DNA Helicase fragment fused to eGFP. Upon phosphorylation by CDKs, this fusion protein changes in localization from the nucleus to the cytoplasm. The combined regulation of transcription and subcellular localization enabled us to visualize the moment of cell cycle entry in dividing seam cells during C. elegans larval development. This reporter is the first to reflect cell cycle commitment in C. elegans and will help further genetic studies of the mechanisms that underlie cell cycle entry and exit.
Highlights
Cell division follows a sequence of events that together result in the segregation of replicated chromosomes and the formation of two new daughter cells
It has long been known that this decision depends on activation of cyclin-dependent kinases (CDKs) in association with G1 cyclins
Live visualization of cell cycle entry in C. elegans cycle entry during development in C. elegans. We show that this marker accurately reflects cell cycle entry and quiescence in divisions of the C. elegans seam cell lineage
Summary
Cell division follows a sequence of events that together result in the segregation of replicated chromosomes and the formation of two new daughter cells. Creating cells in the correct numbers is critical to ensure proper development and tissue homeostasis, while imbalances between the formation and removal of cells within an organism can lead to cancer [1]. The most important decision to determine the creation of cells occurs in the G1 phase, when cells decide whether or not to enter a cell division cycle. It has long been known that this decision depends on activation of cyclin-dependent kinases (CDKs) in association with G1 cyclins. External factors, such as the presence of nutrients (yeasts), growth factors and mitogens (multicellular organisms), determine G1 cyclin expression. Growth factor signalling directly regulates the PLOS ONE | DOI:10.1371/journal.pone.0171600 February 3, 2017
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