Multiple Small RNAs Modulate Rho-Dependent Termination in the Cyclopropane Fatty Acid Synthase mRNA 5’ Untranslated Region

Bacterial small RNAs (sRNAs) have been commonly characterized as regulators of post-transcriptional steps of gene expression including translation and stability of mRNA targets. Previous work revealed that the Escherichia coli cyclopropane fatty acid synthase ( cfa ) mRNA is regulated by at least five different sRNAs by a proposed mechanism involving regulated cfa mRNA turnover by the RNase E degradosome. However, recent work identified the long 5’ untranslated region (UTR) of cfa mRNA as a potential target for Rho-dependent transcription termination, leading us to question whether sRNAs might regulate cfa gene expression at the level of transcription elongation. In this study we report evidence for premature Rho-dependent termination within the long 5’ UTR of cfa , and demonstrate that a pyrimidine-only tract within the 5’ UTR is required for efficient Rho-dependent regulation of cfa . Our data suggest that all of the sequence determinants required for efficient Rho-mediated termination are harbored within the cfa long mRNA 5’ UTR. Finally, we discovered that both the activating sRNA RydC and repressing sRNA CpxQ regulate cfa primarily by modulating Rho-dependent termination of cfa transcription, with only a minor effect on RNase E degradosome-dependent turnover of cfa mRNA. These results illustrate the versatile mechanisms sRNAs use to regulate target gene expression at transcriptional and post-transcriptional levels and suggest that regulation by sRNAs in long UTRs can involve modulation of transcription elongation.