Environmental signal integration by a modular AND gate
J Christopher Anderson1,2, Christopher A Voigt1 & Adam P Arkin2
- Department of Pharmaceutical Chemistry, QB3: California Institute for Quantitative Biological Research, The University of California San Francisco, San Francisco, CA, USA
- Department of Bioengineering, University of California, Howard Hughes Medical Institute, QB3: California Institute for Quantitative Biological Research, Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA
Correspondence to: Christopher A Voigt1 Department of Pharmaceutical Chemistry, The University of California—San Francisco, Box 2540, Room 408C, 1700 4th Street, San Francisco, CA 94158-2330, USA. Tel.: +1 41 55027050; Fax: +1 41 55024690; Email: cavoigt@picasso.ucsf.edu
Received 12 March 2007; Accepted 6 July 2007; Published online 14 August 2007
Article highlights
- Genetic parts are used to construct a near-digital AND gate
- Two input promoters are integrated to activate a single output promoter
- The circuit is modular and can be connected to different inputs and outputs
- A mathematical model is developed that describes how the circuit integrates information
Synopsis
Cells can be 'programmed' by building DNA encoding a series of instructions. Some recent examples include strains of E. coli that have been programmed to record images of light, form two-dimensional patterns, and oscillate like a clock. Logic gates form the core of electronic computing and are an essential component of complex programs. An AND gate integrates two input signals into a single output. If both inputs are ON, then the output is ON. If either or both inputs are OFF, then the output is OFF. AND gates are particularly important to program a bacterium to respond to a microenvironment that is not well defined by a single signal (i.e., high glucose AND low salt). To incorporate a genetic circuit into a larger program, it is critical that it be able to be connected to different inputs and outputs.
Bacteria 'see' their environment using genetic sensors that respond to different stimuli. These sensors can regulate gene expression by activating a promoter. We have constructed a genetic AND gate that uses such promoters as inputs (Figure 1). One promoter leads to the transcription of an mRNA encoding a transcriptional activator. However, stop codons are placed in the activator gene so that the mRNA alone is not sufficient to produce active protein. Only when a tRNA is transcribed from the second input promoter will the activator be produced. Thus, only when the two input promoters are active, the activator will turn on an output promoter. This genetic architecture produces an AND gate with near-digital behavior (Figure 2). Further, we demonstrate that different input promoters, representing different environmental stimuli, can be connected to this circuit.
Figure 1
A schematic representation of the genetic AND gate is shown. Two promoters are the inputs into the gate. The first promoter is linked to the transcription of the amber suppressor tRNA supD. The second promoter drives the transcription of T7 RNA polymerase. The polymerase gene has been modified to contain two amber stop codons (T7ptag). These stop codons are translated as serine when supD is also transcribed. Polymerase is expressed only when both SupD and T7ptag mRNA are present. To characterize the transfer function of the AND gate, two input promoters are used that respond to the small molecules salicylate and arabinose. In addition, the output is connected to the expression of fast-degrading green fluorescent protein.
Full figure and legend (80K)Figures & Tables indexFigure 2
Integration of two inducible promoters by the AND gate. (A) The fluorescence was measured for 64 combinations of inducer in a fluorimeter. The data are shown for (left to right) 0, 3.2 
10-7, 1.3 10-6, 5.2 10-6, 2.1 10-5, 8.3 10-5, 3.3 10-4, and 1.3 10-3 M arabinose, and (bottom to top) 0, 1.5 10-7, 6.1 10-7, 2.4 10-6, 9.8 10-6, 3.9 10-5, 1.6 10-4, and 6.2 10-4 M salicylate. (B) The fluorescence was measured in individual cells using a flow cytometer to determine the population level behavior. The entire population of cells is turned on in the presence of both arabinose and salicylate (1.3 10-3 and 6.2 10-4 M, respectively). When either inducer is not added, the entire population is turned off. There is a 1000-fold induction between the ON and OFF states. The data for this figure were obtained using plasmids pAC-SalSer914, pBACr-AraT7940, and pBR939b (Supplementary information).
Acknowledgements
We thank Sandy Parkinson for supplying plasmid pLC113. JCA is supported by a Damon Runyon Cancer Research Foundation Postdoctoral Fellowship. APA was supported by the Howard Hughes Medical Institute. CAV was supported by the Sloan Foundation, Pew Fellowship, ONR, Packard Fellowship, NIH EY016546, NIH AI067699, NSF BES-0547637, UC-Discovery, and a Sandler Family Opportunity Award. APA, CAV, and JCA are supported by the SynBERC NSF ERC (www.synberc.org).
The authors declare that they have no competing financial interests.
