Abstract
Crosstalk is a major challenge to engineering sophisticated synthetic gene networks. A common approach is to insulate signal-transduction pathways by minimizing molecular-level crosstalk between endogenous and synthetic genetic components, but this strategy can be difficult to apply in the context of complex, natural gene networks and unknown interactions. Here, we show that synthetic gene networks can be engineered to compensate for crosstalk by integrating pathway signals, rather than by pathway insulation. We demonstrate this principle using reactive oxygen species (ROS)-responsive gene circuits in Escherichia coli that exhibit concentration-dependent crosstalk with non-cognate ROS. We quantitatively map the degree of crosstalk and design gene circuits that introduce compensatory crosstalk at the gene network level. The resulting gene network exhibits reduced crosstalk in the sensing of the two different ROS. Our results suggest that simple network motifs that compensate for pathway crosstalk can be used by biological networks to accurately interpret environmental signals.
Highlights
Crosstalk is a major challenge to engineering sophisticated synthetic gene networks
While crosstalk in two-component system signaling pathways often has a negative impact on host fitness, bacteria contain complex regulatory systems that integrate an array of inputs into one output, for example in the sporulation process of Bacillus subtilis[11]
This results in circuits that exhibit reduced crosstalk in the sensing of the two different reactive oxygen species (ROS)
Summary
Crosstalk is a major challenge to engineering sophisticated synthetic gene networks. A common approach is to insulate signal-transduction pathways by minimizing molecular-level crosstalk between endogenous and synthetic genetic components, but this strategy can be difficult to apply in the context of complex, natural gene networks and unknown interactions. In attempts to minimize crosstalk, synthetic biologists have implemented molecular-level insulation by using inputs that are not naturally sensed by host cells[17], by knocking out endogenous genes[18,19], and by mutating and screening for orthogonal genetic components[20,21,22,23] While these strategies have enabled the construction of moderately sized gene networks[24,25], it is an open question as to whether the insulation approach alone will scale to significantly more complex gene networks or will be adaptable to inputs that are sensed by or interface with host cell signaling networks. This results in circuits that exhibit reduced crosstalk in the sensing of the two different ROS
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