Rosenfeld2005 - REPORTS Gene Regulation at the Single-Cell...

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Gene Regulation at the Single-Cell Level Nitzan Rosenfeld, 1 * Jonathan W. Young, 3 Uri Alon, 1 Peter S. Swain, 2 * Michael B. Elowitz 3 . The quantitative relation between transcription factor concentrations and the rate of protein production from downstream genes is central to the function of genetic networks. Here we show that this relation, which we call the gene regulation function (GRF), fluctuates dynamically in individual living cells, thereby limiting the accuracy with which transcriptional genetic circuits can transfer signals. Using fluorescent reporter genes and fusion proteins, we characterized the bacteriophage lambda promoter P R in Escherichia coli .A novel technique based on binomial errors in protein partitioning enabled calibration of in vivo biochemical parameters in molecular units. We found that protein production rates fluctuate over a time scale of about one cell cycle, while intrinsic noise decays rapidly. Thus, biochemical parameters, noise, and slowly varying cellular states together determine the effective single-cell GRF. These results can form a basis for quantitative modeling of natural gene circuits and for design of synthetic ones. The operation of transcriptional genetic cir- cuits ( 1–5 ) is based on the control of pro- moters by transcription factors. The GRF is the relation between the concentration of active transcription factors in a cell and the rate at which their downstream gene products are produced (expressed) through transcrip- tion and translation. The GRF is typically represented as a continuous graph, with the active transcription factor concentration on the x axis and the rate of production of its target gene on the y axis (Fig. 1A). The shape of this function, e.g., the characteristic level of repressor that induces a given response, and the sharpness, or nonlinearity, of this response ( 1 ) determine key features of cellular behavior such as lysogeny switching ( 2 ), developmen- tal cell-fate decisions ( 6 ), and oscillation ( 7 ). Its properties are also crucial for the design of synthetic genetic networks ( 7–11 ). Cur- rent models estimate GRFs from in vitro data ( 12 , 13 ). However, biochemical parame- ters are generally unknown in vivo and could depend on the environment ( 12 ) or cell history ( 14 , 15 ). Moreover, gene regulation may vary from cell to cell or over time. Three funda- mental aspects of the GRF specify the behav- ior of transcriptional circuits at the single-cell level: its mean shape (averaged over many cells), the typical deviation from this mean, and the time scale over which such fluctua- tions persist. Although fast fluctuations should average out quickly, slow ones may introduce errors in the operation of genetic circuits and may pose a fundamental limit on their ac- curacy. In order to address all three aspects, it is necessary to observe gene regulation in in- dividual cells over time.
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Rosenfeld2005 - REPORTS Gene Regulation at the Single-Cell...

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