摘要：Transcriptional repressors provide widespread biological significance in the regulation of gene expression. However, in prokaryotes, it is particularly difficult to find transcriptional repressors that recognize specific target promoters on genome-scale. To address this need, a genetic biosensor for identifying repressors of target promoters was developed in Escherichia coli from a de novo designed genetic circuit. This circuit can convert the negative input of repressors into positive output of reporters, thereby facilitating the selection and identification of repressors. After evaluating the sensitivity and bias, the biosensor was used to identify the repressors of scbA and aco promoters (P scbA and P aco ), which control the transcription of signalling molecule synthase genes in Streptomyces coelicolor and Streptomyces avermitilis, respectively. Two previously unknown repressors of P scbA were identified from a library of TetR family regulators in S. coelicolor , and three novel repressors of P aco were identified from a genomic library of S. avermitilis . Further in vivo and in vitro experiments confirmed that these newly identified repressors attenuated the transcription of their target promoters by direct binding. Overall, the genetic biosensor developed here presents an innovative and powerful strategy that could be applied for identifying genome-wide unknown repressors of promoters in bacteria.
