Abstract
BackgroundAs an intracellular human pathogen, Mycobacterium tuberculosis (Mtb) is facing multiple stressful stimuli inside the macrophage and the granuloma. Understanding Mtb responses to stress is essential to identify new virulence factors and pathways that play a role in the survival of the tubercle bacillus. The main goal of this study was to map the regulatory networks of differentially expressed (DE) transcripts in Mtb upon various forms of genotoxic stress. We exposed Mtb cells to oxidative (H2O2 or paraquat), nitrosative (DETA/NO), or alkylation (MNNG) stress or mitomycin C, inducing double-strand breaks in the DNA. Total RNA was isolated from treated and untreated cells and subjected to high-throughput deep sequencing. The data generated was analysed to identify DE genes encoding mRNAs, non-coding RNAs (ncRNAs), and the genes potentially targeted by ncRNAs.ResultsThe most significant transcriptomic alteration with more than 700 DE genes was seen under nitrosative stress. In addition to genes that belong to the replication, recombination and repair (3R) group, mainly found under mitomycin C stress, we identified DE genes important for bacterial virulence and survival, such as genes of the type VII secretion system (T7SS) and the proline-glutamic acid/proline-proline-glutamic acid (PE/PPE) family. By predicting the structures of hypothetical proteins (HPs) encoded by DE genes, we found that some of these HPs might be involved in mycobacterial genome maintenance. We also applied a state-of-the-art method to predict potential target genes of the identified ncRNAs and found that some of these could regulate several genes that might be directly involved in the response to genotoxic stress.ConclusionsOur study reflects the complexity of the response of Mtb in handling genotoxic stress. In addition to genes involved in genome maintenance, other potential key players, such as the members of the T7SS and PE/PPE gene family, were identified. This plethora of responses is detected not only at the level of DE genes encoding mRNAs but also at the level of ncRNAs and their potential targets.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-3132-1) contains supplementary material, which is available to authorized users.
Highlights
As an intracellular human pathogen, Mycobacterium tuberculosis (Mtb) is facing multiple stressful stimuli inside the macrophage and the granuloma
The genotoxic stresses exerted by H2O2, mitomycin C (MMC) and DETA/Diethylenetriamine/nitric oxide adduct (NO) on Mtb led to more pronounced transcriptomic perturbations with 52, 131 and 723 differentially expressed (DE) genes, respectively (Fig. 1b-d; Additional file 1), while only two DE genes, alkA and ogt were detected under MNNG treatment (Fig. 1e; Additional file 1)
Nitrosative stress in the form of diethylenetriamine nitric oxide adduct (DETA/NO) resulted in the up-regulation of 383 genes and the down-regulation of 340 genes with fold changes ranging from +257 to -27 (Additional file 1)
Summary
As an intracellular human pathogen, Mycobacterium tuberculosis (Mtb) is facing multiple stressful stimuli inside the macrophage and the granuloma. Understanding Mtb responses to stress is essential to identify new virulence factors and pathways that play a role in the survival of the tubercle bacillus. The causative agent of TB, Mycobacterium tuberculosis (Mtb), infects approximately one third of the world’s human population [1]. Mtb experiences endogenous and exogenous oxidative stress [4]. Mtb faces nitrosative stress from reactive nitrogen species (RNS), such as nitric oxide, nitrite, nitrogen dioxide and nitrates [6]. The tubercle bacillus counteracts the effects of ROS and RNS by producing several enzymes, including catalase, peroxidase, superoxide dismutase, and nitrosothiol reductase, to ensure its intracellular survival and persistence [10]
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