Excessive Promoters as Silencers of Genes Horizontally Acquired by Escherichia coli

Abstract

Horizontally acquired genes are usually transcriptionally inactive, although most of them are associated with genomic loci enriched with promoter-like sequences forming "promoter islands." We hypothesized that lateral DNA transfer induces local mutagenesis, accumulating AT base pairs and creating promoter-like sequences, whose occupancy with RNA polymerase and a specific silencer H-NS suppresses the transcription of foreign genes. Error-prone mutagenesis was implemented for the "promoter island" of a foreign gene appY and the promoter region of an inherent gene dps. Derivatives with changed transcriptional activity were selected using a reporter plasmid pET28_eGFP. Only one cycle of mutagenesis with negative selection suppressed the activity of the main dps promoter to the background level due to a single substitution in its-10 element, while positive selection gave a sequence with improved-35 element, thus testifying feasibility of the approach. The same suppression for appY was achieved by three cycles, while eightfold transcription activation required nine iterations of mutagenesis. In both cases, the number of potential start points decreased resulting in an ordinary regulatory region with only one dominant promoter in the case of positive selection. Efficiency of H-NS binding remained virtually unchanged in all mutant constructs. Based on these findings we conclude that excessive promoters can adversely affect transcription by providing a platform for interference between several RNA polymerase molecules, which can act as a silencer at promoter-dense regions.

Escherichia coli transcription factors of unknown function: sequence features and possible evolutionary relationships

“…Recently, experimental evidence of such events has been reported ( Bykov et al, 2020 ) and corroborated the hypothesis that several promoter sequences hinder RNA polymerases’ activity by suppressing the expression of foreign genes. Also, the regions of HGT genes result in accumulating A+T-rich regions and creating promoter-like elements.…”

The emergence and evolution of gene expression in genome regions replete with regulatory motifs

“…We then asked whether gaining a -10 or -35 box motif leads to a significant increase in fluorescence that indicates the creation of a new promoter (Mann-Whitney U-test, Fig. S7 and methods Prompted by an earlier report 18 , we also asked whether mutations that destroy -10 or - 35…”

The LeuO regulator and quiescence: About transcriptional roadblocks, multiple promoters, and crispr‐cas

“…In essence, it may be acting as a genome sentinel by quiescing the expression of foreign-acquired genes that are only needed under very defined physiological conditions. This idea has been put forward recently with the study of two E. coli model genes (Bykov et al, 2020).…”

“…In essence, it may be acting as a genome sentinel by quiescing the expression of foreign‐acquired genes that are only needed under very defined physiological conditions. This idea has been put forward recently with the study of two E. coli model genes (Bykov et al, 2020). One codes for Dps, a NAP, which was considered as the inherent E. coli gene, and the other was appY , coding for a transcriptional regulator and thought to be acquired by horizontal transfer.…”