Identification of Promoter Activity in Gene-Less Cassettes fromVibrionaceaeSuperintegrons

Abstract

ABSTRACTIntegrons are genetic platforms that acquire new genes encoded in integron cassettes (ICs), building arrays of adaptive functions. ICs generally encode promoterless genes, whose expression relies on the platform-associated Pc promoter, with the cassette array functioning as an operon-like structure regulated by the distance to the Pc. This is relevant in large sedentary chromosomal integrons (SCIs) carrying hundreds of ICs, like those inVibriospecies. We selected 29 gene-less cassettes in fourVibrioSCIs, and explored whether their function could be related to the transcription regulation of adjacent ICs. We show that most gene-less cassettes have promoter activity on the sense strand, enhancing the expression of downstream cassettes. Additionally, we identified the transcription start sites of gene-less ICs through 5’-RACE. Accordingly, we found that most of the superintegron inVibrio choleraeis not silent. Thesepromoter cassettescan trigger the expression of a silentdfrB9cassette downstream, increasing trimethoprim resistance >512-fold inV. choleraeandEscherichia coli. Furthermore, one cassette with an antisense promoter can reduce trimethoprim resistance when cloned downstream. Our findings highlight the regulatory role of gene-less cassettes in the expression of adjacent cassettes, emphasizing their significance in SCIs and their clinical importance if captured by mobile integrons.