Rational design of a bacterial transcriptional cascade for amplifying gene expression capacity.
Cascade regulatory circuits have been described that control numerous cell processes, and may provide models for the design of artificial circuits with novel properties. Here we describe the design of a transcriptional regulatory cascade to amplify the cell response to a given signal. We used the salicylate-responsive activators of Pseudomonas putida NahR of the naphthalene degradation plasmid NAH7 and XylS2, a mutant regulator of the TOL plasmid for catabolism of m-xylene and their respective cognate promoters Psal and PM: Control of the expression of xylS2 with the nahR/Psal system permitted either their selective activation with specific effectors for each protein or the simultaneous activation of both of them with salicylate. When cells face the common effector of the two regulators, both the increase in XylS2 concentration and the stimulation of its activity act synergistically on the PM: promoter, amplifying the gene expression capacity by at least one order of magnitude with respect to the individual systems. By changing the hierarchy of regulators, we showed that the specific features of the downstream regulator were crucial for the amplification effect. Directed changes in the effector profile of the regulators allowed the extension of the amplifying system to other molecular signals.
Cebolla
A
A
Departamento de Genética and Departamento de Microbiología y Parasitología, Universidad de Sevilla, 41080 Seville, Spain. acebolla@cica.es
Sousa
C
C
de Lorenzo
V
V
D001426:Bacterial Proteins / Q000235:genetics; D001565:Benzoates; D004274:DNA, Recombinant; D004268:DNA-Binding Proteins; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D007763:Lac Operon / Q000235:genetics; D016958:Pseudomonas putida / Q000187:drug effects / Q000235:genetics*; D012459:Salicylates / Q000494:pharmacology; D015534:Trans-Activators / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects; D001616:beta-Galactosidase / Q000187:drug effects / Q000378:metabolism
D001426:Bacterial Proteins; D001565:Benzoates; D004274:DNA, Recombinant; D004268:DNA-Binding Proteins; C063173:NahR protein, Bacteria; D012459:Salicylates; D015534:Trans-Activators; D014157:Transcription Factors; C078720:XylS protein, Pseudomonas putida; C044605:methyl benzoate; D001616:beta-Galactosidase