Mycobacterium tuberculosis response to cholesterol is integrated with environmental pH and potassium levels via a lipid metabolism regulator

Chen, Yue; MacGilvary, Nathan J.; Tan, Shumin

doi:10.1371/journal.pgen.1011143

Cited by 4 publications

(1 citation statement)

References 69 publications

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“…Mce3R, a TetR-type transcriptional repressor, controls the expression of mel2 genes, including melH ( 11 ). A recent report identified an interrelationship between cholesterol, pH, and potassium levels dependent on Mce3R, emphasizing the crucial role of this regulon in host survival and its importance in responding to environmental stresses within the infected macrophage ( 12 ).…”

Section: Introductionmentioning

confidence: 99%

Uncovering the roles of Mycobacterium tuberculosis melH in redox and bioenergetic homeostasis: implications for antitubercular therapy

Chen,

Yang,

Wang

et al. 2024

mSphere

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Mycobacterium tuberculosis ( Mtb ), the pathogenic bacterium that causes tuberculosis, has evolved sophisticated defense mechanisms to counteract the cytotoxicity of reactive oxygen species (ROS) generated within host macrophages during infection. The melH gene in Mtb and Mycobacterium marinum ( Mm ) plays a crucial role in defense mechanisms against ROS generated during infection. We demonstrate that melH encodes an epoxide hydrolase and contributes to ROS detoxification. Deletion of melH in Mm resulted in a mutant with increased sensitivity to oxidative stress, increased accumulation of aldehyde species, and decreased production of mycothiol and ergothioneine. This heightened vulnerability is attributed to the increased expression of whiB3 , a universal stress sensor. The absence of melH also resulted in reduced intracellular levels of NAD + , NADH, and ATP. Bacterial growth was impaired, even in the absence of external stressors, and the impairment was carbon source dependent. Initial MelH substrate specificity studies demonstrate a preference for epoxides with a single aromatic substituent. Taken together, these results highlight the role of melH in mycobacterial bioenergetic metabolism and provide new insights into the complex interplay between redox homeostasis and generation of reactive aldehyde species in mycobacteria. IMPORTANCE This study unveils the pivotal role played by the melH gene in Mycobacterium tuberculosis and in Mycobacterium marinum in combatting the detrimental impact of oxidative conditions during infection. This investigation revealed notable alterations in the level of cytokinin-associated aldehyde, para -hydroxybenzaldehyde, as well as the redox buffer ergothioneine, upon deletion of melH . Moreover, changes in crucial cofactors responsible for electron transfer highlighted melH ’s crucial function in maintaining a delicate equilibrium of redox and bioenergetic processes. MelH prefers epoxide small substrates with a phenyl substituted substrate. These findings collectively emphasize the potential of melH as an attractive target for the development of novel antitubercular therapies that sensitize mycobacteria to host stress, offering new avenues for combating tuberculosis.

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Section: Introductionmentioning

confidence: 99%

Uncovering the roles of Mycobacterium tuberculosis melH in redox and bioenergetic homeostasis: implications for antitubercular therapy

Chen,

Yang,

Wang

et al. 2024

mSphere

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Mycobacterium tuberculosis Mce3R TetR-like Repressor Forms an Asymmetric Four-Helix Bundle and Binds a Nonpalindrome Sequence

Panagoda,

Balázsi,

Sampson

2024

ACS Chem. Biol.

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Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis, is a major global health concern. TetR family repressors (TFRs) are important for Mtb's adaptation to the human host environment. Our study focuses on one notable Mtb repressor, Mce3R, composed of an unusual double TFR motif. Mce3R-regulated genes encode enzymes implicated in cholesterol metabolism, resistance against reactive oxygen species, and lipid transport activities important for Mtb survival and persistence in the host and for the cellular activity of a 6-azasteroid derivative. Here, we present the structure of Mce3R bound to its DNA operator, unveiling a unique asymmetric assembly previously unreported. We obtained a candidate DNA-binding motif through MEME motif analysis, comparing intergenic regions of mce3R orthologues and identifying nonpalindromic regions conserved between orthologues. Using an electrophoretic mobility shift assay (EMSA), we confirmed that Mce3R binds to a 123-bp sequence that includes the predicted motif. Using scrambled DNA and DNA oligonucleotides of varying lengths with sequences from the upstream region of the yrbE3A (mce3) operon, we elucidated the operator region to be composed of two Mce3R binding sites, each a 25-bp asymmetric sequence separated by 53 bp. Mce3R binds with a higher affinity to the downstream site with a K d of 2.4 ± 0.7 nM. The cryo-EM structure of Mce3R bound to the 123-bp sequence was refined to a resolution of 2.51 Å. Each Mce3R monomer comprises 21 α-helices (α1-α21) folded into an asymmetric TFR-like structure with a core asymmetric four-helix bundle. This complex has two nonidentical HTH motifs and a single ligandbinding domain. The two nonidentical HTHs from each TFR bind within the high-affinity, nonpalindromic operator motif, with Arg53 and Lys262 inserted into the major groove. Site-directed mutagenesis of Arg53 to alanine abrogated DNA binding, validating the Mce3R/DNA structure obtained. Among 811,645 particles, 63% were Mce3R homodimer bound to two duplex oligonucleotides. Mce3R homodimerizes primarily through α15, and each monomer binds to an identical site in the DNA duplex oligonucleotide.

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Uncovering the Roles ofMycobacterium tuberculosis melHin Redox and Bioenergetic Homeostasis: Implications for Antitubercular Therapy

Chen,

Yang,

Wang

et al. 2023

Preprint

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Mycobacterium tuberculosis(Mtb), the pathogenic bacterium that causes tuberculosis, has evolved sophisticated defense mechanisms to counteract the cytotoxicity of reactive oxygen species (ROS) generated within host macrophages during infection. ThemelHgene inMtbandMycobacterium marinum(Mm) plays a crucial role in defense mechanisms against ROS generated during infection. We demonstrate thatmelHencodes an epoxide hydrolase and contributes to ROS detoxification. Deletion ofmelHinMmresulted in a mutant with increased sensitivity to oxidative stress, a minor increase in the bacterium’s sensitivity to NO exposure. This heightened vulnerability is attributed to the increased expression ofwhiB3, subsequently leading to decreased production of mycothiol and ergothioneine. Our study elucidated profound effects on the bioenergetic metabolism ofMmupon disruption ofmelH. The absence ofmelHresulted in reduced intracellular levels of NAD+, NADH, and ATP, and accumulation of aldehydes withinMm.Bacterial growth was impaired, even in the absence of external stressors, and was carbon-source-dependent. Taken together, these results highlight the essential role ofmelHin mycobacterial metabolism and provide new insights into the complex interplay between redox homeostasis and carbon utilization in mycobacteria.ImportanceThis study unveils the pivotal role played by themelHgene inMycobacterium tuberculosisandMycobacterium marinumin combatting the detrimental impact of oxidative conditions during infection. This investigation revealed notable alterations in level of cytokinin-associated aldehyde,para-hydroxybenzaldehyde, as well as the redox buffer ergothioneine, upon deletion ofmelH. Moreover, changes in crucial cofactors responsible for electron transfer highlightedmelH’s crucial function in maintaining a delicate equilibrium of redox and bioenergetic processes. These findings collectively emphasize the potential ofmelHas an attractive target for the development of novel antitubercular therapies, offering new avenues for combating tuberculosis.

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Mycobacterium tuberculosis response to cholesterol is integrated with environmental pH and potassium levels via a lipid metabolism regulator

Cited by 4 publications

References 69 publications

Uncovering the roles of Mycobacterium tuberculosis melH in redox and bioenergetic homeostasis: implications for antitubercular therapy

Uncovering the roles of Mycobacterium tuberculosis melH in redox and bioenergetic homeostasis: implications for antitubercular therapy

Mycobacterium tuberculosis Mce3R TetR-like Repressor Forms an Asymmetric Four-Helix Bundle and Binds a Nonpalindrome Sequence

Uncovering the Roles ofMycobacterium tuberculosis melHin Redox and Bioenergetic Homeostasis: Implications for Antitubercular Therapy

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