Transcriptome sequencing of the human pathogen Corynebacterium diphtheriae NCTC 13129 provides detailed insights into its transcriptional landscape and into DtxR-mediated transcriptional regulation

Abstract

BackgroundThe human pathogen Corynebacterium diphtheriae is the causative agent of diphtheria. In the 1990s a large diphtheria outbreak in Eastern Europe was caused by the strain C. diphtheriae NCTC 13129. Although the genome was sequenced more than a decade ago, not much is known about its transcriptome. Our aim was to use transcriptome sequencing (RNA-Seq) to close this knowledge gap and gain insights into the transcriptional landscape of a C. diphtheriae tox+ strain.ResultsWe applied two different RNA-Seq techniques, one to retrieve 5′-ends of primary transcripts and the other to characterize the whole transcriptional landscape in order to gain insights into various features of the C. diphtheriae NCTC 13129 transcriptome. By examining the data we identified 1656 transcription start sites (TSS), of which 1202 were assigned to genes and 454 to putative novel transcripts. By using the TSS data promoter regions recognized by the housekeeping sigma factor σA and its motifs were analyzed in detail, revealing a well conserved −10 but an only weakly conserved −35 motif, respectively. Furthermore, with the TSS data 5’-UTR lengths were explored. The observed 5’-UTRs range from zero length (leaderless transcripts), which make up 20% of all genes, up to over 450 nt long leaders, which may harbor regulatory functions. The C. diphtheriae transcriptome consists of 471 operons which are further divided into 167 sub-operon structures. In a differential expression analysis approach, we discovered that genetic disruption of the iron-sensing transcription regulator DtxR, which controls expression of diphtheria toxin (DT), causes a strong influence on general gene expression. Nearly 15% of the genome is differentially transcribed, indicating that DtxR might have other regulatory functions in addition to regulation of iron metabolism and DT. Furthermore, our findings shed light on the transcriptional landscape of the DT encoding gene tox and present evidence for two tox antisense RNAs, which point to a new way of transcriptional regulation of toxin production.ConclusionsThis study presents extensive insights into the transcriptome of C. diphtheriae and provides a basis for future studies regarding gene characterization, transcriptional regulatory networks, and regulation of the tox gene in particular.Electronic supplementary materialThe online version of this article (10.1186/s12864-018-4481-8) contains supplementary material, which is available to authorized users.

Defining the Transcriptional and Post-transcriptional Landscapes of Mycobacterium smegmatis in Aerobic Growth and Hypoxia

“…The apparent robustness of translation is shared with Mtb, where 25% of the transcripts lack a leader sequence (Cortes et al, 2013; Shell et al, 2015b). In addition, high abundances of transcripts lacking 5′ UTRs have been reported in other bacteria including Corynebacterium diphtheria, Leptospira interrogans, Borrelia burgdorferi , and Deinococcus deserti , the latter having 60% leaderless transcripts (de Groot et al, 2014; Adams et al, 2017; Zhukova et al, 2017; Wittchen et al, 2018). Considering the high proportion of leaderless transcripts and the large number of leadered transcripts that lack a SD sequence (53%), it follows that an important number of transcripts are translated without canonical interactions between the mRNA and anti-Shine-Dalgarno sequence, suggesting that M. smegmatis has versatile mechanisms to address translation.…”

Overlap of Promoter Recognition Specificity of Stress Response Sigma Factors SigD and SigH in Corynebacterium glutamicum ATCC 13032

“…Transcription start sites (TSS) were detected essentially as described by Wittchen et al (2018). Primary 5′-end cDNA library data were analyzed with the software ReadXplorer v2.2 (Hilker et al, 2016) using the Transcription analysis function.…”

NGS-based phylogeny of diphtheria-related pathogenicity factors in different Corynebacterium spp. implies species-specific virulence transmission

“…However, it is not known yet if surrounding prophages or PAIs are absolutely species-specific or also transmitted between Corynebacterium species. DT is furthermore part of an iron-dependent regulation system with the diphtheria toxin repressor (DtxR) acting as main regulatory control center on DT, but also on various other virulence factors [16]. In contrast to the DT encoding tox gene, the dtxR gene occurs both in toxigenic and non-toxigenic isolates of the different Corynebacterium species and its genetic variability between them has not been analysed in detail yet.…”