A cis-RNA-mediated expression regulation of the tet(M) resistance gene in Enterococcus faecium

Abstract

A set of putative novel small RNAs was recently identified as expressed in Enterococcus faecium, a major opportunistic pathogen involved in numerous healthcare-associated infections and hospital outbreaks. The aim of this study was to characterize the first functional analysis of one of them, srn0030, by phenotypic, genomic and transcriptomic approaches. By genomic analysis and RACE mapping, we revealed the presence of this RNA (previously designated as Ptet) within the 5-untrasnlated region (UTR) of tet(M), a gene conferring tetracycline resistance through ribosomal protection. The regulatory mechanism has previously been described as transcriptional attenuation, but has actually been poorly characterized. Hence, we provide original additional data, especially the presence of three upstream transcripts of ~100, ~150 and ~230 nt within the 5-UTR of tet(M), suggesting an alternative regulatory mechanism. The total deletion of these three transcripts causes an unexpected decreasing of tetracycline resistance in E. faecium. The attenuation mechanism was investigated, and we confirmed that the transcriptional read-through and tet(M) overexpression induced by tetracycline addition but the function of putative peptide leader on attenuation mechanism was not supported by our data. We report here new phenotypic and transcriptomic observations in E. faecium demonstrating an alternative regulatory mechanism of tet(M) gene expression.