Abstract
The behavior of genetic circuits is often poorly predictable. A gene’s expression level is not only determined by the intended regulators, but also affected by changes in ribosome availability imparted by expression of other genes. Here we design a quasi-integral biomolecular feedback controller that enables the expression level of any gene of interest (GOI) to adapt to changes in available ribosomes. The feedback is implemented through a synthetic small RNA (sRNA) that silences the GOI’s mRNA, and uses orthogonal extracytoplasmic function (ECF) sigma factor to sense the GOI’s translation and to actuate sRNA transcription. Without the controller, the expression level of the GOI is reduced by 50% when a resource competitor is activated. With the controller, by contrast, gene expression level is practically unaffected by the competitor. This feedback controller allows adaptation of genetic modules to variable ribosome demand and thus aids modular construction of complicated circuits.
Highlights
The behavior of genetic circuits is often poorly predictable
Modularity, the property according to which the i/o behavior of a system does not change with its context, is required for bottomup design of synthetic genetic circuits[33]
Modular design of genetic circuits relies on the assumption that the i/o behavior of TX devices is not affected by surrounding systems
Summary
The behavior of genetic circuits is often poorly predictable. A gene’s expression level is determined by the intended regulators, and affected by changes in ribosome availability imparted by expression of other genes. Activation of one transcriptional (TX) device, that is, a system where input TX regulators affect expression level of one output protein, reduces availability of ribosomes to other, otherwise unconnected, TX devices, affecting their gene-expression levels by up to 60%7,11. These unintended nonregulatory interactions among TX devices can significantly alter the emergent behavior of genetic circuits. Feedback control has played a critical role in making circuit’s components modular, i.e., in maintaining a desired i/o response despite disturbances This enabled predictable and reliable composition of larger systems from subsystems[14]. A promising proof of concept, the requirement to co-design the TX device’s input and output to engineer the feedback prevents generalization and scalability of this solution beyond the specific circuit’s instance considered
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