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Mycobacterium tuberculosis transcriptional response to small molecule

ABSTRACT: We applied RNAseq (Nextseq) technology to study mechanism of action of nitro-containing heterocycle antitubercular JSF-2019, with des-nitro JSF-2026 as control. Briefly, mid-log phase (OD = 0.3) M. tuberculosis culture was treated by 10x MIC of each compound in biological quadruplicates followed by mRNA extraction and RNAseq analysis. We find that JSF-2019 and pretomanid as intracellular NO• donors exhibited distinct transcriptional patterns comparing to extracellular iNOS inducer such as DETA/NO, therefore it suggests distinguished mechanisms of action between intracellular vs. extracellular NO• donors. We also observed that JSF-2019 upregulated a subset of FAS-II genes similar to isoniazid, indicating JSF-2019 inhibits intracellular mycolic acid biosynthesis.

ORGANISM(S): Mycobacterium tuberculosis

PROVIDER: GSE126718 | GEO | 2019/02/19

REPOSITORIES: GEO

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Project description:Mycobacterium tuberculosis has a complex cell envelope that is remodelled throughout infection to respond and survive the hostile and variable intracellular conditions within the host. Despite the importance of cell wall homeostasis in pathogenicity, little is known about the environmental signals and regulatory networks controlling cell wall biogenesis in mycobacteria. The mycolic acid desaturase regulator (MadR) is a transcriptional repressor responsible for regulation of the essential aerobic desaturases desA1 and desA2 that are differentially regulated throughout infection along with mycolate modification genes and thus, likely involved in mycolic acid remodelling. Here we generated a madR null mutant in M. smegmatis that exhibited traits of an impaired cell wall with increased permeability, susceptibility to rifampicin and cell surface disruption as a consequence of desA1/desA2 dysregulation. Analysis of mycolic acids revealed the presence of a highly desaturated mycolate in the null mutant that exists in relative trace amounts in the wildtype, but increases in abundance upon cell surface disruption as a result of relieved repression on the desA1/desA2 promoters. Transcriptomic profiling confirmed MadR as a cell surface disruption responsive regulator of desA1/desA2 and further implicating it in the control of bespoke β-oxidation pathways and transport evolutionarily diversified subnetworks associated with virulence. In vitro characterisation of MadR using electromobility shift assays and analysis of binding affinities is suggestive of a unique acyl-CoA pool sensing mechanism, whereby MadR is able to bind a range of acyl-CoA but MadR repression of desA1/desA2 promoters is only relieved upon binding of saturated acyl-CoA of chain length C16-C24. We propose this acyl effector ligand mechanism as distinct to other regulators of mycolic acid biosynthesis or fatty acid desaturases and places MadR as the key regulatory checkpoint that coordinates mycolic acid remodelling in response to host derived cell surface perturbation

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Mycobacterium tuberculosis transcriptional response to small molecule

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