Small RNA MTS1338 Configures a Stress Resistance Signature in Mycobacterium tuberculosis

Abstract

In the course of evolution, Mycobacterium tuberculosis (Mtb), the etiological agent of tuberculosis, has developed sophisticated strategies to evade host immune response, including the synthesis of small non-coding RNAs (sRNAs), which regulate post-transcriptional pathways involved in the stress adaptation of mycobacteria. sRNA MTS1338 is upregulated in Mtb during its infection of cultured macrophages and in the model of chronic tuberculosis, suggesting involvement in host-pathogen interactions. Here, we analyzed the role of MTS1338 in the Mtb response to macrophage-like stresses in vitro. The Mtb strain overexpressing MTS1338 demonstrated enhanced survival ability under low pH, nitrosative, and oxidative stress conditions simulating the antimicrobial environment inside macrophages. Transcriptomic analysis revealed that in MTS1338-overexpressing Mtb, the stress factors led to the activation of a number of transcriptional regulators, toxin-antitoxin modules, and stress chaperones, about half of which coincided with the genes induced in Mtb phagocytosed by macrophages. We determined the MTS1338 "core regulon", consisting of 11 genes that were activated in all conditions under MTS1338 overexpression. Our findings indicate that MTS1338 is a stress-induced sRNA that promotes Mtb survival in macrophages by triggering adaptive transcriptional mechanisms in response to host antimicrobial defense reactions.

Keywords: Mycobacterium tuberculosis; RNA-seq; pathogen; small non-coding RNA; stress resistance; survival; transcriptomic signature.

Figure 1

MTS1338 overexpression promotes Mtb survival in infected THP-1 cells. At 4 h post infection (p.i.), non-phagocytosed bacteria were removed, then the infected THP-1 cells were incubated for 20 h (24 h p.i.) and 44 h (48 h p.i.). CFU values were measured in triplicate and are presented as the mean ± SD; * p < 0.05, ** p < 0.01.

Figure 2

MTS1338 overexpression in Mtb promotes stress tolerance. Stress effects were measured by using [³H]-uracil incorporation; the viability of mycobacteria not subjected to stresses was taken as 100%. The data are presented as the mean ± SD; * p < 0.05.

Figure 3

Two-component principal component analysis (PCA) of MTS1338-over (circles) and pMV-empty (triangles) samples under various conditions.

Figure 4

Transcriptional differences between MTS1338-overexpressing and pMV-empty strains in stressful and ‘no stress’ (control) conditions. Unfilled circles indicate genes with log₂FC ≤ 1.5 and/or padj > 0.1, filled circles—genes with log₂FC > 1.5 and padj < 0.1, red circles—condition-specific DEGs, and green circles—common genes in all the conditions tested.

Figure 5

Relationships among DEGs in the MTS1338-overexpressing strain exposed to different conditions.