Conditional termination of transcription is shaped by Rho and translated uORFS inMycobacterium tuberculosis

Abstract

Accurately controlled termination of transcription is critical for gene expression and adaptation to changing environments. The aim of this study was to define transcription termination sites in Mycobacterium tuberculosis at nucleotide resolution. To this end, we applied Term-seq to cultures of M. tuberculosis and established that by far the most abundant mechanism of transcription termination is Rho-dependent. Moreover, we find that conditional or premature termination of transcription regulates a large proportion of genes and that this is often associated with translated upstream open reading frames (uORFs). Finally, our results show that overlapping ORFs are abundant in many cases in the absence of a recognisable Shine-Dalgarno sequence, indicating tightly coordinated/coupled translation. Our results provide a detailed image and comprehensive catalogue of novel M. tuberculosis cis-regulatory elements, where Rho-dependent conditional termination of transcription and translational coupling together play major roles in gene expression control.

Structural and functional basis of the universal transcription factor NusG pro-pausing activity inMycobacterium tuberculosis

“…Further, their biological functions may differ. The existence of translationally coupled-attenuation mechanisms suggests transcription and translation are coupled in Mtb (D’Halluin et al, 2022; Lee et al, 2022) as they are in the model g-proteobacterium Eco where NusG mediates transcription–translation coupling (Burmann et al, 2010). Bsu transcription is uncoupled from translation (Johnson et al, 2020) and, unlike for Mtb (Figure 6), Bsu NusG is dispensable for growth.…”

Using a Whole Genome Co-expression Network to Inform the Functional Characterisation of Predicted Genomic Elements fromMycobacterium tuberculosisTranscriptomic Data

“…Ribosome profiling (Ribo-seq) methods that sequence the ribosome-protected fragments of mRNA have identified actively translated RNA in the 5' UTRs of annotated protein-coding mRNA transcripts (Canestrari et al, 2020;D'Halluin et al, 2022;Sawyer et al, 2021;Shell et al, 2015;C. Smith et al, 2022).…”

“…Though RNA-seq-based ncRNA prediction algorithms are often assumed to overpredict putative ncRNAs, especially at the 5’ and 3’ ends of coding genes, there are biological and technical reasons for detecting abundant signal in the unannotated regions of the genome. Ribosome profiling (Ribo-seq) methods that sequence the ribosome-protected fragments of mRNA have identified actively translated RNA in the 5’ UTRs of annotated protein-coding mRNA transcripts (Canestrari et al, 2020; D’Halluin et al, 2022; Sawyer et al, 2021; Shell et al, 2015; C. Smith et al, 2022). These unannotated sORFs may represent functional peptides or function to regulate the translation of the downstream transcript; however, it is impossible to tell the difference between a putative ncRNA and a sORF from RNA-seq signal alone.…”

“…Ribosome profiling (Ribo-seq) methods that sequence the ribosome-protected fragments of mRNA have identified actively translated RNA in the 5' UTRs of annotated protein-coding mRNA transcripts (Canestrari et al, 2020;D'Halluin et al, 2022;Sawyer et al, 2021;Shell et al, 2015;Smith et al, 2022). These unannotated sORFs may represent functional peptides or function to regulate the translation of the downstream transcript; however, it is impossible to tell the difference between a putative ncRNA and a sORF from RNA-seq signal alone.…”

“…These unannotated sORFs may represent functional peptides or function to regulate the translation of the downstream transcript; however, it is impossible to tell the difference between a putative ncRNA and a sORF from RNA-seq signal alone. Additionally, the 3’ ends in mycobacterial RNA-seq data often lack clear signal termination (Dar et al, 2016; D’Halluin et al, 2022; Lejars et al, 2019) and processing of transcripts at the 3’ end may be the norm (Wang et al, 2019). Finally, polycistronic transcripts often include non-coding sequence between the genes of an operon, and this may contain functional elements and/or processing sites (Martini et al, 2019).…”