The MarR-Type Regulator PA3458 Is Involved in Osmoadaptation Control in Pseudomonas aeruginosa

Kotecka, Karolina; Kawałek, Adam; Kobyłecki, Kamil; Bartosik, Aneta Agnieszka

doi:10.3390/ijms22083982

Cited by 14 publications

(14 citation statements)

References 90 publications

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“…3D-E) (Table S3). This result is surprising, as in other bacteria, including E. coli (single MarR regulator) and P. aeruginosa (13 MarR regulators), loss of MarR activity confers resistance to a diversity of antibiotics (53)(54)(55).…”

Section: Transposon Insertions In Marr Confer a Growth Advantage In T...mentioning

confidence: 95%

“…These data suggest that ETH may have the DmarR did not exhibit cross resistance to other antibiotics tested (Table S3), suggesting a unique function in mediating ETH resistance. This was surprising, as MarR homologs often confer resistance to multiple antibiotics in other bacterial species (53)(54)(55). The fact that resistant mutants fail to emerge in the combinatorial treatment of ETH with AMK and CLR suggests that there may be unique mechanisms of resistance to each of these antibiotics.…”

Section: Eth Treatment With Clr or Amk Suppresses The Emergence Of Re...mentioning

confidence: 99%

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MarR-Dependent Transcriptional Regulation of mmpSL5 induces Ethionamide Resistance in Mycobacterium abscessus

Rodriguez¹,

Campbell-Kruger

Camba

et al. 2022

Preprint

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Mycobacterium abscessus (Mabs) is an emerging non-tuberculosis mycobacterial (NTM) pathogen responsible for a wide variety of respiratory and cutaneous infections that are difficult to treat with standard antibacterial therapy. Mabs has a high degree of both innate and acquired antibiotic resistance to most clinically relevant drugs, including standard anti-mycobacterial agents. Ethionamide (ETH), an inhibitor of mycolic acid biosynthesis is currently utilized as a second-line agent for treating multidrug resistant tuberculosis (MDR-TB) infections. Here, we show that ETH has activity against clinical strains of Mabs in vitro at concentrations that are therapeutically achievable. Using transposon mutagenesis and whole genome sequencing of spontaneous drug-resistant mutants, we identified marR (MAB_2648c) as a genetic determinant of ETH sensitivity in Mabs. The gene marR encodes a transcriptional regulator of the TetR family of regulators. We show that MarR represses expression of MAB_2649 (mmpS5) and MAB_2650 (mmpL5). Further, we show that de-repression of these genes in marR mutants confers resistance to ETH, but not other antibiotics. To identify determinants of resistance that may be shared across antibiotics, we also performed Tn-Seq during treatment with amikacin and clarithromycin, drugs currently used clinically to treat Mabs. We found very little overlap in genes that modulate the sensitivity of Mabs to all three antibiotics, suggesting a high degree of specificity for resistance mechanisms in this emerging pathogen.

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Section: Transposon Insertions In Marr Confer a Growth Advantage In T...mentioning

confidence: 95%

Section: Eth Treatment With Clr or Amk Suppresses The Emergence Of Re...mentioning

confidence: 99%

MarR-Dependent Transcriptional Regulation of mmpSL5 induces Ethionamide Resistance in Mycobacterium abscessus

Rodriguez¹,

Campbell-Kruger

Camba

et al. 2022

Preprint

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“…The oligomerization state of purified His 6 -PA3027 was assayed by crosslinking as described previously [84]. 4.6.…”

Section: Glutaraldehyde Crosslinkingmentioning

confidence: 99%

The AraC-Type Transcriptional Regulator GliR (PA3027) Activates Genes of Glycerolipid Metabolism in Pseudomonas aeruginosa

Kotecka

Kawałek

Kobyłecki

et al. 2021

IJMS

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Pseudomonas aeruginosa encodes a large set of transcriptional regulators (TRs) that modulate and manage cellular metabolism to survive in variable environmental conditions including that of the human body. The AraC family regulators are an abundant group of TRs in bacteria, mostly acting as gene expression activators, controlling diverse cellular functions (e.g., carbon metabolism, stress response, and virulence). The PA3027 protein from P. aeruginosa has been classified in silico as a putative AraC-type TR. Transcriptional profiling of P. aeruginosa PAO1161 overexpressing PA3027 revealed a spectacular increase in the mRNA levels of PA3026-PA3024 (divergent to PA3027), PA3464, and PA3342 genes encoding proteins potentially involved in glycerolipid metabolism. Concomitantly, chromatin immunoprecipitation-sequencing (ChIP-seq) analysis revealed that at least 22 regions are bound by PA3027 in the PAO1161 genome. These encompass promoter regions of PA3026, PA3464, and PA3342, showing the major increase in expression in response to PA3027 excess. In Vitro DNA binding assay confirmed interactions of PA3027 with these regions. Furthermore, promoter-reporter assays in a heterologous host showed the PA3027-dependent activation of the promoter of the PA3026-PA3024 operon. Two motifs representing the preferred binding sites for PA3027, one localized upstream and one overlapping with the −35 promoter sequence, were identified in PA3026p and our data indicate that both motifs are required for full activation of this promoter by PA3027. Overall, the presented data show that PA3027 acts as a transcriptional regulator in P. aeruginosa, activating genes likely engaged in glycerolipid metabolism. The GliR name, from a glycerolipid metabolism regulator, is proposed for PA3027 of P. aeruginosa.

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“…To standardize the assays, all plates were prepared with the same volume of the medium. Biofilm amount was measured with the crystal violet staining method in a rich medium [79].…”

Section: Motility and Biofilm Formation Assaysmentioning

confidence: 99%

“…Strains obtained by transformation of PAO1161 with pMEB12 or pKGB8 were used for total RNA preparation. RNA isolation and sequencing as well as data analysis were performed essentially as previously described [79]. Raw data are available in the NCBI's Gene Expression Omnibus (GEO) database (http://www.ncbi.nlm.nih.gov/geo/ (accessed on 30 October 2021)) under accession number GSE186749.…”

Section: Rna Isolation Rna-seq and Rt-qpcrmentioning

confidence: 99%

The Lrp/AsnC-Type Regulator PA2577 Controls the EamA-like Transporter Gene PA2576 in Pseudomonas aeruginosa

Modrzejewska

Kawałek

Bartosik

2021

IJMS

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The regulatory network of gene expression in Pseudomonas aeruginosa, an opportunistic human pathogen, is very complex. In the PAO1 reference strain, about 10% of genes encode transcriptional regulators, many of which have undefined regulons and unknown functions. The aim of this study is the characterization of PA2577 protein, a representative of the Lrp/AsnC family of transcriptional regulators. This family encompasses proteins involved in the amino acid metabolism, regulation of transport processes or cell morphogenesis. The transcriptome profiling of P. aeruginosa cells with mild PA2577 overproduction revealed a decreased expression of the PA2576 gene oriented divergently to PA2577 and encoding an EamA-like transporter. A gene expression analysis showed a higher mRNA level of PA2576 in P. aeruginosa ΔPA2577, indicating that PA2577 acts as a repressor. Concomitantly, ChIP-seq and EMSA assays confirmed strong interactions of PA2577 with the PA2577/PA2576 intergenic region. Additionally, phenotype microarray analyses indicated an impaired metabolism of ΔPA2576 and ΔPA2577 mutants in the presence of polymyxin B, which suggests disturbances of membrane functions in these mutants. We show that PA2576 interacts with two proteins, PA5006 and PA3694, with a predicted role in lipopolysaccharide (LPS) and membrane biogenesis. Overall, our results indicate that PA2577 acts as a repressor of the PA2576 gene coding for the EamA-like transporter and may play a role in the modulation of the cellular response to stress conditions, including antimicrobial peptides, e.g., polymyxin B.

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The MarR-Type Regulator PA3458 Is Involved in Osmoadaptation Control in Pseudomonas aeruginosa

Cited by 14 publications

References 90 publications

MarR-Dependent Transcriptional Regulation of mmpSL5 induces Ethionamide Resistance in Mycobacterium abscessus

MarR-Dependent Transcriptional Regulation of mmpSL5 induces Ethionamide Resistance in Mycobacterium abscessus

The AraC-Type Transcriptional Regulator GliR (PA3027) Activates Genes of Glycerolipid Metabolism in Pseudomonas aeruginosa

The Lrp/AsnC-Type Regulator PA2577 Controls the EamA-like Transporter Gene PA2576 in Pseudomonas aeruginosa

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