“Tight control of cell cycle entry is essential both during normal development and in tissue homeostasis.”
The majority of cells in the adult body are not proliferating and a fraction of these cells are quiescent. Quiescence is a state of reversible cell cycle arrest, from which cells can be stimulated to re-enter the cell cycle. During tissue maintenance and repair, the transition from quiescence to proliferation and proliferation to quiescence must be carefully regulated to maintain tissue size and function. Aberrant regulation of these cell cycle control mechanisms can drive the continuous proliferation of cells, promoting tumorigenesis.
Our aim is to understand how proliferation-quiescence decisions are made at the molecular level. We investigate how these systems are controlled in normal human cells, and then determine how mutations and aberrations that occur in tumour cells can drive their continuous proliferation. One approach that we use to interrogate cell cycle control systems is quantitative, live, single-cell imaging of CRISPR-engineered cells. This is a powerful technique that allows us to measure the real-time expression of signalling proteins as cells undergo quiescence-proliferation transitions and thus determine how the coordinated output of this dynamical cell system regulates cellular phenotype.
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A. CRISPR-engineering of gene loci to introduce fluorescent proteins into proteins of interest. B. Sample imaging data – single cell shown over time. C. Sample data from live imaging experiment – each curve is p21-GFP levels in a single cell shown from mitotic exit.
Alexis Barr holds a Cancer Research UK Career Development Fellowship.
Stojic L, Lun ATL, Mangei J, Mascalchi P, Quarantotti V, Barr AR, Bakal C, Marioni J, Gergely F, Odom DT. (2018). Specificity of RNAi, LNA and CRISPRi as loss-of-function methods in transcriptional analysis. Nucleic Acids Res doi: 10.1093/nar/gky437.
Heldt FS, Barr AR, Cooper S, Bakal C, Novak B. (2018). A comprehensive model for the proliferation-quiescence decision in response to endogenous DNA damage in human cells. PNAS doi: 10.1073/pnas.1715345115.
Asghar US, Barr AR, Cutts R, Beaney M, Babina I, Sampath D, Giltnane J, Arca Lacap J, Crocker L, Young A, Pearson A, Herrera-Abreu MT, Bakal C, Turner NC. (2017). Single-cell dynamics determines response to CDK4/6 inhibition in triple negative breast cancer. Clinical Cancer Research 23(18), 5561-5572. doi: 10.1158/1078-0432.CCR-17-0369.
Barr AR, Cooper S, Heldt FS, Butera F, Stoy H, Mansfeld J, Novak B, Bakal C. (2017). DNA damage during S-phase mediates the proliferation-quiescence decision in the subsequent G1 via p21 expression. Nature Communications 8, 14728. doi: 10.1038/ncomms14728.
Barr AR, Heldt FS, Zhang T, Bakal C, Novak B. (2016). A dynamical framework for the all-or-none G1/S transition. Cell Systems 2(1), 27-37. doi: 10.1016/j.cels.2016.01.001.