Natural RNA Polymerase Aptamers Regulate Transcription in E. coli

Abstract

Summary In search for RNA signals that modulate transcription via direct interaction with RNA polymerase (RNAP) we deep-sequenced an E. coli genomic library enriched for RNAP-binding RNAs. Many natural RNAP-binding aptamers, termed RAPs, were mapped to the genome. Over 60% of E. coli genes carry RAPs in their mRNA. Combining in vitro and in vivo approaches we characterized a subset of RAPs (iRAPs) that promote Rho-dependent transcription termination. A representative iRAP within the coding region of the essential gene, nadD, greatly reduces its transcriptional output in stationary phase and under oxidative stress, demonstrating that iRAPs control gene expression in response to changing environment. The mechanism of iRAPs involves active uncoupling of transcription and translation, making nascent RNA accessible to Rho. iRAPs encoded in the antisense strand also promote gene expression by reducing transcriptional interference. In essence, our work uncovers a broad class of cis-acting RNA signals that globally control bacterial transcription.

Aptamers: Uptake mechanisms and intracellular applications

“…The majority of identified RAPs (64.3%) was characterized inhibitory RAPs (iRAPs) that encoded in the strand opposite to the annotated genes acting like transcriptional 'anti-interference'. The antisense iRAPs curbed transcriptional interference by cis-acting as pro-termination signals, potentiating the expression of sense-encoded genes [43]. This study shows that natural RNA aptamers have potential as a self-regulatory system.…”

“…Conventionally, artificial aptamers are isolated against their targets with a synthetic aptamer library pool. To systematically search for natural RNA aptamers that regulate transcription through direct interaction with RNA polymerase (RNAP) in bacteria, genomic SELEX; a genomic library of E. coli as a source of RNA sequences and the E. coli RNAP holoenzyme as bait, were performed [43]. Following the nextgeneration sequencing and functional analysis, 15,000 of natural RNA aptamers to RNAP (RAPs) were identified in the E. coli genome.…”

Ultrasensitive molecular controllers for quasi-integral feedback

“…The switch stage has been recently demonstrated in vitro [52]. While at the moment only two viral RNA polymerases (SP6, T7) have been regulated using aptamers, it is possible to select aptamers against other polymerases or transcription factors using SELEX [72]; natural aptamers regulating RNA polymerase in E. coli have been recently discovered [73].…”

DNA breaks-mediated cost reveals RNase HI as a new target for selectively eliminating antibiotic resistance

“…S1, Table S1). Erythromycin targets the 50S ribosomal subunit, affecting translation and its coupling with transcription (20).…”

“…This allowed us to confirm that resistant mutants indeed show increased DSBs (Extended Data Figure 1, Extended Data Table 1). Erythromycin targets the 50S ribosomal subunit, affecting translation and its coupling with transcription 21 . Erythromycin resistance mutations (Erm R ) mapping in the genes rplD and rplV (encoding L4 and L22, respectively) are known to reduce translation elongation rate 22 , credibly affecting transcription-translation coupling as well.…”