The Spencer lab focuses on the question of when and how cells commit to the cell cycle. Historically, cell cycle commitment was studied by bulk biochemical analysis. Mitogen-starved cells were re-stimulated with mitogens for varying amounts of time to identify a ‘restriction point’ when the presence of mitogens is no longer necessary to complete the cell cycle. This work suggested that the restriction point occurred in the late G1 phase of the cell cycle, concomitant with an upregulation of cyclin dependent kinase 2 (CDK2) activity. However, it was not clear whether this model held true for proliferating populations, nor was the mechanism behind the restriction point understood.
To tackle these questions, we developed a live-cell sensor to monitor CDK2 activity in individual cycling cells. The sensor consists of a YFP-tagged section of DNA helicase B (DHB) with four CDK phosphorylation sites close to a nuclear localization sequence (NLS) and a nuclear export sequence (NES) (Fig. 1A). Phosphorylation of the sensor by CDK2 unmasks the NES and causes translocation of the sensor to the cytoplasm in proportion to CDK2 activity. By monitoring the cytoplasmic:nuclear ratio of this sensor as a readout for CDK2 activity, we discovered a bifurcation point at mitotic exit where cells choose between two future fates. Some newly born cells immediately build up CDK2 activity and commit to the next cell cycle (CDK2inc cells, Fig. 1B blue), while others lack CDK2 activity and enter a G0-like state (CDK2low cells, Fig. 1B red). We showed that this bifurcation is directly controlled by the CDK inhibitor p21 and is regulated by mitogens during a ‘restriction window’ (RW1) at the end of the previous cell cycle. Cells in the CDK2low state face a second mitogen-sensitive window (RW2, the classic restriction point) in which they can choose to build up CDK2 activity to re-enter the cell cycle (Fig. 1C).
Following these results, the current focus of the lab is to understand how diverse pathways, from mitogenic signaling to DNA damage, are integrated to control cell cycle entry in both normal and cancer cells. Projects in the lab range from basic regulation of the cell cycle, to sensor and tool development, to more translational projects looking at the misregulated proliferation of cancer cells. We are actively recruiting new members, so if you would like to hear more about specific projects in the lab, please contact Sabrina Spencer.