TY - JOUR
T1 - Measurement of gene regulation in individual cells reveals rapid switching between promoter states
AU - Sepúlveda, Leonardo A.
AU - Xu, Heng
AU - Zhang, Jing
AU - Wang, Mengyu
AU - Golding, Ido
N1 - Funding Information:
We are grateful to the following people for generous advice and for providing reagents: I. Dodd, M. Elowitz, L. Finzi, H. Garcia, T. Gregor, T. Kuhlman, L. McLane, R. Phillips, A. Raj, E. Rothenberg, A. Sanchez, K. Shearwin, R. Singer, S. Skinner, L-H. So, A. Sokac, L. Zeng, and C. Zong. Work in the Golding lab is supported by grants from NIH (R01 GM082837), NSF (PHY 1147498, PHY 1430124 and PHY 1427654), The Welch Foundation (Q-1759), and The John S. Dunn Foundation (Collaborative Research Award). H.X. is supported by the Burroughs Wellcome Fund Career Award at the Scientific Interface. We gratefully acknowledge the computing resources provided by the Computational and Integrative Biomedical Research Center of Baylor College of Medicine.
PY - 2016/3/11
Y1 - 2016/3/11
N2 - In vivo mapping of transcription-factor binding to the transcriptional output of the regulated gene is hindered by probabilistic promoter occupancy, the presence of multiple gene copies, and cell-To-cell variability.We demonstrate how to overcome these obstacles in the lysogeny maintenance promoter of bacteriophage lambda, PRM. We simultaneously measured the concentration of the lambda repressor CI and the number of messenger RNAs (mRNAs) from PRM in individual Escherichia coli cells, and used a theoretical model to identify the stochastic activity corresponding to different CI binding configurations. We found that switching between promoter configurations is faster than mRNA lifetime and that individual gene copies within the same cell act independently. The simultaneous quantification of transcription factor and promoter activity, followed by stochastic theoretical analysis, provides a tool that can be applied to other genetic circuits.
AB - In vivo mapping of transcription-factor binding to the transcriptional output of the regulated gene is hindered by probabilistic promoter occupancy, the presence of multiple gene copies, and cell-To-cell variability.We demonstrate how to overcome these obstacles in the lysogeny maintenance promoter of bacteriophage lambda, PRM. We simultaneously measured the concentration of the lambda repressor CI and the number of messenger RNAs (mRNAs) from PRM in individual Escherichia coli cells, and used a theoretical model to identify the stochastic activity corresponding to different CI binding configurations. We found that switching between promoter configurations is faster than mRNA lifetime and that individual gene copies within the same cell act independently. The simultaneous quantification of transcription factor and promoter activity, followed by stochastic theoretical analysis, provides a tool that can be applied to other genetic circuits.
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U2 - 10.1126/science.aad0635
DO - 10.1126/science.aad0635
M3 - Article
C2 - 26965629
AN - SCOPUS:84960921360
VL - 351
SP - 1218
EP - 1222
JO - Science
JF - Science
SN - 0036-8075
IS - 6278
ER -