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Origin of bistability underlying mammalian cell cycle entry.

Yao G, Tan C, West M, Nevins JR, You L - Mol. Syst. Biol. (2011)

Bottom Line: We identified a minimal circuit that is able to generate robust, resettable bistability.Underscoring its importance, experimental disruption of this circuit abolishes maintenance of the activated E2F state, supporting its importance for the bistability of the Rb-E2F system.Our findings suggested basic design principles for the robust control of the bistable cell cycle entry at the R-point.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular & Cellular Biology, University of Arizona, Tucson, AZ 85721, USA. guangyao@arizona.edu

ABSTRACT
Precise control of cell proliferation is fundamental to tissue homeostasis and differentiation. Mammalian cells commit to proliferation at the restriction point (R-point). It has long been recognized that the R-point is tightly regulated by the Rb-E2F signaling pathway. Our recent work has further demonstrated that this regulation is mediated by a bistable switch mechanism. Nevertheless, the essential regulatory features in the Rb-E2F pathway that create this switching property have not been defined. Here we analyzed a library of gene circuits comprising all possible link combinations in a simplified Rb-E2F network. We identified a minimal circuit that is able to generate robust, resettable bistability. This minimal circuit contains a feed-forward loop coupled with a mutual-inhibition feedback loop, which forms an AND-gate control of the E2F activation. Underscoring its importance, experimental disruption of this circuit abolishes maintenance of the activated E2F state, supporting its importance for the bistability of the Rb-E2F system. Our findings suggested basic design principles for the robust control of the bistable cell cycle entry at the R-point.

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Experimental disruption of the gene circuit 3–5–6–7 abolishes the Rb–E2F bistable switch. (A) The Cdk2 inhibitor CVT-313 selectively blocks the mutual-inhibition feedback loop 5–6, but not the other three positive-feedback loops (9, 2–3–6, and 2–7). (B) Experimental protocol of serum-pulse stimulation and Cdk2 inhibition. At time 0, cells were serum-starved and at quiescence. See Materials and methods for details. (C) The influence of Cdk2 inhibition on E2F bistability. Each curve represents the histogram of the E2F–d2GFP distribution from approximately 5000 cells. The Cdk2 inhibitor dose and sample harvest time are as indicated. The dashed lines connecting the high and low E2F–d2GFP modes are for the guide of eyes.
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f3: Experimental disruption of the gene circuit 3–5–6–7 abolishes the Rb–E2F bistable switch. (A) The Cdk2 inhibitor CVT-313 selectively blocks the mutual-inhibition feedback loop 5–6, but not the other three positive-feedback loops (9, 2–3–6, and 2–7). (B) Experimental protocol of serum-pulse stimulation and Cdk2 inhibition. At time 0, cells were serum-starved and at quiescence. See Materials and methods for details. (C) The influence of Cdk2 inhibition on E2F bistability. Each curve represents the histogram of the E2F–d2GFP distribution from approximately 5000 cells. The Cdk2 inhibitor dose and sample harvest time are as indicated. The dashed lines connecting the high and low E2F–d2GFP modes are for the guide of eyes.

Mentions: To this end, we constructed and analyzed a library of mathematical models that encompass all possible circuit designs derived from a simplified Rb–E2F network (Figure 1). We identified a minimal gene circuit that is uniquely robust in generating resettable bistability. This minimal circuit consists of a mutual-inhibition feedback loop between the Rb (RP) and E2F (EE) modules (Figure 1, links 5, 6) and a feed-forward loop between the Myc (MD) and E2F (EE) modules (Figure 1, links 7, 3, 6). These two regulatory motifs form an AND-gate control of E2F activation (system output). Our modeling analysis suggested that the mutual-inhibition feedback loop between the Rb and E2F modules is critical for generating a robust bistable switch. Meanwhile, the feed-forward loop between the Myc and E2F modules and the AND-gate control are critical for the resettability. Underscoring the importance of this model-predicted minimal circuit, targeted disruption of this circuit abolishes maintenance of the activated E2F state, supporting its necessity for the bistability of the Rb–E2F system (Figure 3).


Origin of bistability underlying mammalian cell cycle entry.

Yao G, Tan C, West M, Nevins JR, You L - Mol. Syst. Biol. (2011)

Experimental disruption of the gene circuit 3–5–6–7 abolishes the Rb–E2F bistable switch. (A) The Cdk2 inhibitor CVT-313 selectively blocks the mutual-inhibition feedback loop 5–6, but not the other three positive-feedback loops (9, 2–3–6, and 2–7). (B) Experimental protocol of serum-pulse stimulation and Cdk2 inhibition. At time 0, cells were serum-starved and at quiescence. See Materials and methods for details. (C) The influence of Cdk2 inhibition on E2F bistability. Each curve represents the histogram of the E2F–d2GFP distribution from approximately 5000 cells. The Cdk2 inhibitor dose and sample harvest time are as indicated. The dashed lines connecting the high and low E2F–d2GFP modes are for the guide of eyes.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3101952&req=5

f3: Experimental disruption of the gene circuit 3–5–6–7 abolishes the Rb–E2F bistable switch. (A) The Cdk2 inhibitor CVT-313 selectively blocks the mutual-inhibition feedback loop 5–6, but not the other three positive-feedback loops (9, 2–3–6, and 2–7). (B) Experimental protocol of serum-pulse stimulation and Cdk2 inhibition. At time 0, cells were serum-starved and at quiescence. See Materials and methods for details. (C) The influence of Cdk2 inhibition on E2F bistability. Each curve represents the histogram of the E2F–d2GFP distribution from approximately 5000 cells. The Cdk2 inhibitor dose and sample harvest time are as indicated. The dashed lines connecting the high and low E2F–d2GFP modes are for the guide of eyes.
Mentions: To this end, we constructed and analyzed a library of mathematical models that encompass all possible circuit designs derived from a simplified Rb–E2F network (Figure 1). We identified a minimal gene circuit that is uniquely robust in generating resettable bistability. This minimal circuit consists of a mutual-inhibition feedback loop between the Rb (RP) and E2F (EE) modules (Figure 1, links 5, 6) and a feed-forward loop between the Myc (MD) and E2F (EE) modules (Figure 1, links 7, 3, 6). These two regulatory motifs form an AND-gate control of E2F activation (system output). Our modeling analysis suggested that the mutual-inhibition feedback loop between the Rb and E2F modules is critical for generating a robust bistable switch. Meanwhile, the feed-forward loop between the Myc and E2F modules and the AND-gate control are critical for the resettability. Underscoring the importance of this model-predicted minimal circuit, targeted disruption of this circuit abolishes maintenance of the activated E2F state, supporting its necessity for the bistability of the Rb–E2F system (Figure 3).

Bottom Line: We identified a minimal circuit that is able to generate robust, resettable bistability.Underscoring its importance, experimental disruption of this circuit abolishes maintenance of the activated E2F state, supporting its importance for the bistability of the Rb-E2F system.Our findings suggested basic design principles for the robust control of the bistable cell cycle entry at the R-point.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular & Cellular Biology, University of Arizona, Tucson, AZ 85721, USA. guangyao@arizona.edu

ABSTRACT
Precise control of cell proliferation is fundamental to tissue homeostasis and differentiation. Mammalian cells commit to proliferation at the restriction point (R-point). It has long been recognized that the R-point is tightly regulated by the Rb-E2F signaling pathway. Our recent work has further demonstrated that this regulation is mediated by a bistable switch mechanism. Nevertheless, the essential regulatory features in the Rb-E2F pathway that create this switching property have not been defined. Here we analyzed a library of gene circuits comprising all possible link combinations in a simplified Rb-E2F network. We identified a minimal circuit that is able to generate robust, resettable bistability. This minimal circuit contains a feed-forward loop coupled with a mutual-inhibition feedback loop, which forms an AND-gate control of the E2F activation. Underscoring its importance, experimental disruption of this circuit abolishes maintenance of the activated E2F state, supporting its importance for the bistability of the Rb-E2F system. Our findings suggested basic design principles for the robust control of the bistable cell cycle entry at the R-point.

Show MeSH