Cytoplasmic RNA Sensor Pathways and Nitazoxanide Broadly Inhibit Intracellular Mycobacterium tuberculosis Growth

Abstract

SummaryTo establish stable infection, Mycobacterium tuberculosis (MTb) must overcome host innate immune mechanisms, including those that sense pathogen-derived nucleic acids. Here, we show that the host cytosolic RNA sensing molecules RIG-I-like receptor (RLR) signaling proteins RIG-I and MDA5, their common adaptor protein MAVS, and the RNA-dependent kinase PKR each independently inhibit MTb growth in human cells. Furthermore, we show that MTb broadly stimulates RIG-I, MDA5, MAVS, and PKR gene expression and their biological activities. We also show that the oral FDA-approved drug nitazoxanide (NTZ) significantly inhibits intracellular MTb growth and amplifies MTb-stimulated RNA sensor gene expression and activity. This study establishes prototypic cytoplasmic RNA sensors as innate restriction factors for MTb growth in human cells and it shows that targeting this pathway is a potential host-directed approach to treat tuberculosis disease.

RNA Sensing of Mycobacterium tuberculosis and Its Impact on TB Vaccination Strategies

“…This induction was observed only upon stimulation with live but not dead bacteria. Furthermore, it was shown that activation of RIG-I and MDA5 inhibited mycobacterial growth in different cell lines [60]. There is also evidence for a synergy of the DNA-recognizing cyclic GMP-AMP synthase (cGAS)/stimulator of interferon genes (STING)-axis with the MDA5-IRF7 pathway, activated by cytosolic RNA secreted from the endosome leading to IFNβ expression [61].…”

“…The role of PKR, for which it was shown that sensing mycobacterial dsRNA leads to inhibition of translation, apoptosis, and ultimately decreased mycobacterial growth, is somewhat controversial as it has been recently questioned by retrospective investigation [65]: The observed difference between assumed KO mice in the PKR domain was attributed to different mice strains rather than to the impact of PKR itself. Nevertheless, recent cell line experiments could demonstrate that PKR activation by mycobacterial RNA leads to growth restriction in human cells [60].…”

“…In summary, cytosolic RNA sensing in TB immunity is a currently expanding research field with mounting evidence for its significance for pathogenesis. This may be of importance with regard to new therapeutic options, since for example the drug Nitazoxanide seems to activate this pathway leading to mycobacterial growth inhibition [60].…”

“…At the same time, activation of TLR8 enhances production of reactive oxygen species (ROS) through activation of Cytochrome b (-245) beta (CYBB)/NADPH oxidase 2 (NOX2), which will themselves activate the canonical inflammation pathway through Cathepsin B translocating to the cytosol [52,73]. (b) After transition to the cytosol, mycobacterial RNA stimulates NLRP3 of the inflammasome, which lead to caspase-1 dependent production of IL-18, IL-1β and pyroptosis [50,51], as well as the RLRs Rig-1 and MDA5, leading to expression of type I IFNs [60]. These cytokines will again stimulate NK cells to produce IFNγ.…”

“…As a result, virulence between MTB and BCG differs significantly, as well as the triggered immunological pathways and memory response. With regard to RNA sensing, cytosolic activation of RLRs, PKR, and NLRP3 will thus not happen, but these receptors were shown to be able to contribute to mycobacterial growth restriction [60]. It was shown that an rBCG strain, which, in contrast to canonical BCG, has access to cytosolic presentation of antigens, leads to superior protection against TB correlating with higher levels of central TfH memory cells in mice [122] (Figure 3, pathway b).…”