Genetic circuits in living cells share transcriptional and translational resources that are available in limited amounts. This leads to unexpected couplings among seemingly unconnected modules, which result in poorly predictable circuit behavior. In this study, we determine these interdependencies between products of different genes by characterizing the economy of how transcriptional and translational resources are allocated to the production of proteins in genetic circuits. We discover that, when expressed from the same plasmid, the combinations of attainable protein concentrations are constrained by a linear relationship, which can be interpreted as an isocost line, a concept used in microeconomics. We created a library of circuits with two reporter genes, one constitutive and the other inducible in the same plasmid, without a regulatory path between them. In agreement with the model predictions, experiments reveal that the isocost line rotates when changing the ribosome binding site strength of the inducible gene and shifts when modifying the plasmid copy number. These results demonstrate that isocost lines can be employed to predict how genetic circuits become coupled when sharing resources and provide design guidelines for minimizing the effects of such couplings.

Summary: Microbial communities are increasingly utilized in biotechnology. Efficiency and productivity in many of these applications depends on the presence of cooperative interactions between members of the community. Two key processes underlying these interactions are the production of public goods and metabolic cross‐feeding, which can be understood in the general framework of ecological and evolutionary (eco‐evo) dynamics. In this review, we illustrate the relevance of cooperative interactions in microbial biotechnological processes, discuss their mechanistic origins and analyse their evolutionary resilience. Cooperative behaviours can be damaged by the emergence of ‘cheating’ cells that benefit from the cooperative interactions but do not contribute to them. Despite this, cooperative interactions can be stabilized by spatial segregation, by the presence of feedbacks between the evolutionary dynamics and the ecology of the community, by the role of regulatory systems coupled to the environmental conditions and by the action of horizontal gene transfer. Cooperative interactions enrich microbial communities with a higher degree of robustness against environmental stress and can facilitate the evolution of more complex traits. Therefore, the evolutionary resilience of microbial communities and their ability to constraint detrimental mutants should be considered to design robust biotechnological applications.

Significance: Evolution by natural selection is driven by fitness differences, which define a “fitness landscape” in the space of all possible genetic sequences. Understanding the landscape is critical for understanding and predicting natural selection, yet very little is known about the structure of a real fitness landscape. Here we experimentally determine the complete fitness landscape of small RNA selected to interact with GTP, a building block for early life. We find that the landscape is composed of largely disconnected islands of active sequences. This scenario suggests that natural selection under these conditions would be constrained to local exploration of sequence space but that replaying the initial emergence of a functional RNA could lead to a different outcome.