A Novel TetR-Like Transcriptional Regulator Is Induced in Acid-Nitrosative Stress and Controls Expression of an Efflux Pump in Mycobacteria

Perrone, Filomena; Siena, Barbara De; Muscariello, Lidia; Kendall, Sharon L.; Waddell, Simon J.; Sacco, Margherita

doi:10.3389/fmicb.2017.02039

Cited by 15 publications

(20 citation statements)

References 44 publications

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“…Furthermore, we identified a few putative transcription factors that might be implicated in the regulation of efflux pumps under acid stress (like DeoR, RTCIAT899_RS29995; TetR, RTCIAT899_RS10760, RTCIAT899_RS28830, or YebC/PmpR, RTCIAT899_RS14585). The TetR family of transcriptional regulators has been reported to be involved in the regulation of efflux pumps (Perrone et al, 2017 ) and drug efflux in mycobacteria (Betts et al, 2003 ; Wei et al, 2014 ). Among the DEGs that were up-regulated were also the hyc genes encoding hydrogenases proteins.…”

Section: Discussionmentioning

confidence: 99%

Dissecting the Acid Stress Response of Rhizobium tropici CIAT 899

2018

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Rhizobium tropici CIAT899 is a nodule-forming α-proteobacterium displaying intrinsic resistance to several abiotic stress conditions such as low pH and high temperatures, which are common in tropical environments. It is a good competitor for Phaseolus vulgaris (common bean) nodule occupancy at low pH values, however little is known about the genetic and physiological basis of the tolerance to acidic conditions. To identify genes in R. tropici involved in pH stress response we combined two different approaches: (1) A Tn5 mutant library of R. tropici CIAT899 was screened and 26 acid-sensitive mutants were identified. For 17 of these mutants, the transposon insertion sites could be identified. (2) We also studied the transcriptomes of cells grown under different pH conditions using RNA-Seq. RNA was extracted from cells grown for several generations in minimal medium at 6.8 or 4.5 (adapted cells). In addition, we acid-shocked cells pre-grown at pH 6.8 for 45 min at pH 4.5. Of the 6,289 protein-coding genes annotated in the genome of R. tropici CIAT 899, 383 were differentially expressed under acidic conditions (pH 4.5) vs. control condition (pH 6.8). Three hundred and fifty one genes were induced and 32 genes were repressed; only 11 genes were induced upon acid shock. The acid stress response of R. tropici CIAT899 is versatile: we found genes encoding response regulators and membrane transporters, enzymes involved in amino acid and carbohydrate metabolism and proton extrusion, in addition to several hypothetical genes. Our findings enhance our understanding of the core genes that are important during the acid stress response in R. tropici.

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Section: Discussionmentioning

confidence: 99%

Dissecting the Acid Stress Response of Rhizobium tropici CIAT 899

2018

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“…By GFP promoter probe and transcriptional analyses, we strengthened that observation and defined the TetR regulons. We also showed, by EMSA analysis, that the recombinant purified TetR MSMEG-3765 protein is able to bind both MSMEG_3762 and Rv1687c promoters, indicating binding affinity of the M. smegmatis protein for the Mtb regulatory region ( Perrone et al, 2017 ). This, together with the high percentage of identity between the two operons, suggests that the two efflux systems may work in an analogous way in Mtb and M. smegmatis .…”

Section: Introductionmentioning

confidence: 75%

“…Members of the TetR family of transcriptional regulators are widespread among bacteria, especially in microorganisms exposed to environmental stresses, being involved in the control of many cellular functions, including efflux pump activity ( Balhana et al, 2015 ). We have previously demonstrated that in M. smegmatis , the TetR-like protein MSMEG-3765 (TetR3765) regulates expression of its own operon, coding for two other proteins annotated as components of an ABC efflux pump, and that the recombinant purified TetR3765 protein is able to bind both MSMEG_3762/63/65 and Rv1687/86/85c upstream sequences, indicating binding affinity of the M. smegmatis protein for the Mtb regulatory region ( Perrone et al, 2017 ). These results suggested that the same regulatory system exists for expression of the putative MSMEG-3762/63 and Rv1687/86c efflux pumps, which share a high level of amino acid sequence identity.…”

Section: Discussionmentioning

confidence: 99%

Characterization of the Mycobacterial MSMEG-3762/63 Efflux Pump in Mycobacterium smegmatis Drug Efflux

et al. 2020

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Multi-drug resistant tuberculosis (MDR-TB) represents a major health problem worldwide. Drug efflux and the activity of efflux transporters likely play important roles in the development of drug-tolerant and drug-resistant mycobacterial phenotypes. This study is focused on the action of a mycobacterial efflux pump as a mechanism of drug resistance. Previous studies demonstrated up-regulation of the TetR-like transcriptional regulator MSMEG_3765 in Mycobacterium smegmatis and its ortholog Rv1685c in Mycobacterium tuberculosis (Mtb) in acid-nitrosative stress conditions. MSMEG-3765 regulates the expression of the MSMEG_3762/63/65 operon, and of the orthologous region in Mtb (Rv1687c/86c/85c). MSMEG-3762 and Rv1687c are annotated as ATP-binding proteins, while MSMEG-3763 and Rv1686c are annotated as trans-membrane polypeptides, defining an ABC efflux pump in both M. smegmatis and Mtb. The two putative efflux systems share a high percentage of identity. To examine the role of the putative efflux system MSMEG-3762/63, we constructed and characterized a MSMEG-3763 deletion mutant in M. smegmatis (∆MSMEG_3763). By comparative analysis of wild type, knockout, and complemented strains, together with structural modeling and molecular docking bioinformatics analyses of the MSMEG-3763 trans-membrane protein, we define the protein complex MSMEG-3762/63 as an efflux pump. Moreover, we demonstrate involvement of this pump in biofilm development and in the extrusion of rifampicin and ciprofloxacin (CIP), antimicrobial drugs used in first- and second-line anti-TB therapies.

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“…3A, lanes 4–6). Similar results were obtained using a reported nucleoid‐associated protein, NapM, but not the transcriptional regulator MSMEG_3765 (Liu et al ., 2016; Perrone et al ., 2017) (Fig. 3A, lanes 7–12).…”

Section: Resultsmentioning

confidence: 98%

“…S4B). The MSMEG_3765 transcription regulator bound its promoter, but not to Ms2227p (Perrone et al ., 2017) (Fig. S4C).…”

Section: Resultsmentioning

confidence: 99%

MmbR, a master transcription regulator that controls fatty acid β‐oxidation genes in Mycolicibacterium smegmatis

蘇

et al. 2020

Environmental Microbiology

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Summary An unusually high lipid content and a complex lipid profile are the most distinctive features of the mycobacterial cell envelope. However, our understanding of the regulatory mechanism underlying mycobacterial lipid metabolism is limited, and the major regulators responsible for lipid homeostasis remain to be characterized. Here, we identified MmbR as a novel master regulator that is essential for maintaining lipid homeostasis in Mycolicibacterium smegmatis. We found that MmbR controls fatty acid β‐oxidation and modulates biofilm formation in Mycolicibacterium smegmatis. Although MmbR possesses the properties of nucleoid‐associated proteins, it acts as a TetR‐like transcription factor, directly regulating and intensively repressing the expression of a group of core genes involved in fatty acid β‐oxidation. Furthermore, both long‐chain acyl‐Coenzyme A and fatty acids appear to regulate the signal molecules modulated by MmbR. The deletion of mmbR led to a significant reduction in intracellular fatty acid content and a decrease in the relative lipid composition of the biofilm. The lack of mmbR led to morphological changes in the mycobacterial colony, defects in biofilm formation and enhanced sensitivity to anti‐tuberculosis drugs. Our study is the first to establish a link between the transcriptional regulation of fatty acid β‐oxidation genes and lipid homeostasis in mycobacteria.

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A Novel TetR-Like Transcriptional Regulator Is Induced in Acid-Nitrosative Stress and Controls Expression of an Efflux Pump in Mycobacteria

Cited by 15 publications

References 44 publications

Dissecting the Acid Stress Response of Rhizobium tropici CIAT 899

Dissecting the Acid Stress Response of Rhizobium tropici CIAT 899

Characterization of the Mycobacterial MSMEG-3762/63 Efflux Pump in Mycobacterium smegmatis Drug Efflux

MmbR, a master transcription regulator that controls fatty acid β‐oxidation genes in Mycolicibacterium smegmatis

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