scholarly journals Adaptation of Cupriavidus metallidurans CH34 to Toxic Zinc Concentrations Involves an Uncharacterized ABC-Type Transporter

2021 ◽  
Vol 9 (2) ◽  
pp. 309
Author(s):  
Rob Van Houdt ◽  
Joachim Vandecraen ◽  
Natalie Leys ◽  
Pieter Monsieurs ◽  
Abram Aertsen
Keyword(s):  
Abc Transporter ◽  
Rhizobium Leguminosarum ◽  
Constitutive Expression ◽  
Metal Resistance ◽  
Cadmium Resistance ◽  
Gene Encoding ◽  
Cupriavidus Metallidurans Ch34 ◽  
Cupriavidus Metallidurans ◽  
High Zinc ◽  
Zinc Concentrations

Cupriavidus metallidurans CH34 is a well-studied metal-resistant β-proteobacterium and contains a battery of genes participating in metal metabolism and resistance. Here, we generated a mutant (CH34ZnR) adapted to high zinc concentrations in order to study how CH34 could adaptively further increase its resistance against this metal. Characterization of CH34ZnR revealed that it was also more resistant to cadmium, and that it incurred seven insertion sequence-mediated mutations. Among these, an IS1088 disruption of the glpR gene (encoding a DeoR-type transcriptional repressor) resulted in the constitutive expression of the neighboring ATP-binding cassette (ABC)-type transporter. GlpR and the adjacent ABC transporter are highly similar to the glycerol operon regulator and ATP-driven glycerol importer of Rhizobium leguminosarum bv. viciae VF39, respectively. Deletion of glpR or the ABC transporter and complementation of CH34ZnR with the parental glpR gene further demonstrated that loss of GlpR function and concomitant derepression of the adjacent ABC transporter is pivotal for the observed resistance phenotype. Importantly, addition of glycerol, presumably by glycerol-mediated attenuation of GlpR activity, also promoted increased zinc and cadmium resistance in the parental CH34 strain. Upregulation of this ABC-type transporter is therefore proposed as a new adaptation route towards metal resistance.

scholarly journals The Response of Cupriavidus Metallidurans CH34 to Cadmium Involves a Decrease in Intracellular Levels of C-Di-GMP Probably Mediated by a Novel Metal Regulated Phosphodiesterase That Inhibits the Biofilm Lifestyle

2018 ◽  
Author(s):  
Pablo Alviz ◽  
Sebastian Fuentes ◽  
Luis Rojas ◽  
Raymond Turner ◽  
Michael Seeger ◽  
...  
Keyword(s):  
Heavy Metal ◽  
Biofilm Formation ◽  
Bioinformatic Analysis ◽  
Heavy Metal Resistance ◽  
Metal Resistance ◽  
Cadmium Resistance ◽  
Toxic Heavy Metal ◽  
Transcription Analysis ◽  
Cupriavidus Metallidurans Ch34 ◽  
Cupriavidus Metallidurans

Cadmium is a highly toxic heavy metal for biological systems. Cupriavidus metallidurans CH34 is a model strain for heavy metal resistance and bioremediation. The aim of this study was to determine the role of the c-di-GMP pathway in the C. metallidurans CH34 response to cadmium in both planktonic and biofilm cells. Increasing cadmium concentrations correlates with an inhibition of biofilm formation and EPS production in C. metallidurans cells. Planktonic and biofilm cells showed similar tolerance to cadmium. During exposure to cadmium an acute decrease of c-di-GMP levels in planktonic and biofilm cells was observed. Transcription analysis by RT-qPCR showed that cadmium induced in planktonic cells and strongly induced in biofilm cells the expression of the urf2 gene and the mercuric reductase encoding merA gene, which belong to the Tn501/Tn21 mer operon. After exposure to cadmium the cadA gene involved in cadmium resistance was equally upregulated in both lifestyles. Bioinformatic analysis and null mutant complementation assays indicated that the protein encoded by the urf2 gene is a functional phosphodiesterase involved in the c-di-GMP metabolism. We propose to rename the urf2 gene as mrp gene for metal regulated phosphodiesterase. An increase of the second messenger c-di-GMP content by the heterologous expression of the constitutively active diguanylate cyclase PleD* correlated with an increase in biofilm formation and cadmium susceptibility. These results indicate that the response to cadmium in C. metallidurans CH34 involves a decrease in c-di-GMP content that inhibits the biofilm lifestyle.

scholarly journals The Components of the Unique Zur Regulon of Cupriavidus metallidurans Mediate Cytoplasmic Zinc Handling

2017 ◽  
Vol 199 (21) ◽  
Author(s):  
Lucy Bütof ◽  
Christopher Schmidt-Vogler ◽  
Martin Herzberg ◽  
Cornelia Große ◽  
Dietrich H. Nies
Keyword(s):  
Consensus Sequence ◽  
Regulatory Region ◽  
Metal Resistance ◽  
Resistant Bacterium ◽  
Zinc Homeostasis ◽  
Mineral Salts ◽  
Content Type ◽  
Zip Family ◽  
Cupriavidus Metallidurans ◽  
Zinc Concentrations

ABSTRACT Zinc is an essential trace element, yet it is toxic at high concentrations. In the betaproteobacterium Cupriavidus metallidurans, the highly efficient removal of surplus zinc from the periplasm is responsible for the outstanding metal resistance of the organism. Rather than having a typical Zur-dependent, high-affinity ATP-binding cassette transporter of the ABC protein superfamily for zinc uptake at low concentrations, C. metallidurans has the secondary zinc importer ZupT of the zinc-regulated transporter, iron-regulated transporter (ZRT/IRT)-like protein (ZIP) family. It is important to understand, therefore, how this zinc-resistant bacterium copes with exposure to low zinc concentrations. Members of the Zur regulon in C. metallidurans were identified by comparing the transcriptomes of a Δzur mutant and its parent strain. The consensus sequence of the Zur-binding box was derived for the zupTp promoter-regulatory region by use of a truncation assay. The motif was used to predict possible Zur boxes upstream of Zur regulon members. The binding of Zur to these boxes was confirmed. Two Zur boxes upstream of the cobW 1 gene, encoding a putative zinc chaperone, proved to be required for complete repression of cobW 1 and its downstream genes in cells cultivated in mineral salts medium. A Zur box upstream of each of zur-cobW 2, cobW 3, and zupT permitted both low expression levels of these genes and their upregulation under conditions of zinc starvation. This demonstrates a compartmentalization of zinc homeostasis in C. metallidurans, where the periplasm is responsible for the removal of surplus zinc, cytoplasmic components are responsible for the management of zinc as an essential cofactor, and the two compartments are connected by ZupT. IMPORTANCE Elucidating zinc homeostasis is necessary for understanding both host-pathogen interactions and the performance of free-living bacteria in their natural environments. Escherichia coli acquires zinc under conditions of low zinc concentrations via the Zur-controlled ZnuABC importer of the ABC superfamily, and this was also the paradigm for other bacteria. In contrast, the heavy-metal-resistant bacterium C. metallidurans achieves high tolerance to zinc through sophisticated zinc handling and efflux systems operating on periplasmic zinc ions, so that removal of surplus zinc is a periplasmic feature in this bacterium. It is shown here that this process is augmented by the management of zinc by cytoplasmic zinc chaperones, whose synthesis is controlled by the Zur regulator. This demonstrates a new mechanism, involving compartmentalization, for organizing zinc homeostasis.

scholarly journals Behind the shield of Czc: ZntR controls expression of the gene for the zinc-exporting P-type ATPase ZntA in Cupriavidus metallidurans

2021 ◽  
Author(s):  
Vladislava Schulz ◽  
Christopher Schmidt-Vogler ◽  
Phillip Strohmeyer ◽  
Stefanie Weber ◽  
Daniel Kleemann ◽  
...  
Keyword(s):  
Cadmium Concentration ◽  
Zinc Content ◽  
Metal Resistance ◽  
Zinc Uptake ◽  
Uptake System ◽  
Zinc Ions ◽  
Cupriavidus Metallidurans ◽  
Zinc Concentrations ◽  
Main Regulator ◽  
Cellular Zinc

In the metallophilic beta-proteobacterium Cupriavidus metallidurans, the plasmid-encoded Czc metal homeostasis system adjusts the periplasmic zinc, cobalt and cadmium concentration, which influences subsequent uptake of these metals into the cytoplasm. Behind this shield, the PIB2-type APTase ZntA is responsible for removal of surplus cytoplasmic zinc ions, thereby providing a second level of defense against toxic zinc concentrations. ZntA is the counterpart to the Zur-regulated zinc uptake system ZupT and other import systems; however, the regulator of zntA expression was unknown. The chromid-encoded zntA gene is adjacent to the genes czcI2C2B2’, which are located on the complementary DNA strand and transcribed from a common promoter region. These genes encode homologs of plasmid pMOL30-encoded Czc components. Candidates for possible regulators of zntA were identified and subsequently tested: CzcI, CzcI2, and the MerR-type gene products of the locus tags Rmet_2302, Rmet_0102, Rmet_3456. This led to the identification of Rmet_3456 as ZntR, the main regulator of zntA expression. Moreover, both CzcIs decreased Czc-mediated metal resistance, possibly to avoid “over-excretion” of periplasmic zinc ions, which could result in zinc starvation due to diminished zinc uptake into the cytoplasm. Rmet_2302 was identified as CadR, the regulator of the cadA gene for an important cadmium-exporting PIB2-type ATPase, which provides another system for removal of cytoplasmic zinc and cadmium. Rmet_0102 was not involved in regulation of the metal resistance systems examined here. Thus, ZntR forms a complex regulatory network with CadR, Zur and the CzcIs. Moreover, these discriminating regulatory proteins assign the efflux systems to their particular function. Importance Zinc is an essential metal for numerous organisms from humans to bacteria. The transportome of zinc uptake and efflux systems controls the overall cellular composition and zinc content in a double feed-back loop. Zinc starvation mediates, via the Zur regulator, an up-regulation of the zinc import capacity via the ZIP-type zinc importer ZupT and an amplification of zinc storage capacity, which together raise the cellular zinc content again. On the other hand, an increasing zinc content leads to ZntR-mediated up-regulation of the zinc efflux system ZntA, which decreases the zinc content. Together, the Zur regulon components and ZntR/ZntA balance the cellular zinc content under both high external zinc concentrations and zinc starvation conditions.

scholarly journals The Transcriptomic Landscape of Cupriavidus metallidurans CH34 Acutely Exposed to Copper

Genes ◽  
2020 ◽  
Vol 11 (9) ◽  
pp. 1049
Author(s):  
Laurens Maertens ◽  
Natalie Leys ◽  
Jean-Yves Matroule ◽  
Rob Van Houdt
Keyword(s):  
Sulfur Metabolism ◽  
Model Organism ◽  
Gene Clusters ◽  
Resistance Mechanisms ◽  
Metal Resistance ◽  
Copper Resistance ◽  
Messenger Rnas ◽  
Antisense Transcripts ◽  
Cupriavidus Metallidurans Ch34 ◽  
Cupriavidus Metallidurans

Bacteria are increasingly used for biotechnological applications such as bioremediation, biorecovery, bioproduction, and biosensing. The development of strains suited for such applications requires a thorough understanding of their behavior, with a key role for their transcriptomic landscape. We present a thorough analysis of the transcriptome of Cupriavidus metallidurans CH34 cells acutely exposed to copper by tagRNA-sequencing. C. metallidurans CH34 is a model organism for metal resistance, and its potential as a biosensor and candidate for metal bioremediation has been demonstrated in multiple studies. Several metabolic pathways were impacted by Cu exposure, and a broad spectrum of metal resistance mechanisms, not limited to copper-specific clusters, was overexpressed. In addition, several gene clusters involved in the oxidative stress response and the cysteine-sulfur metabolism were induced. In total, 7500 transcription start sites (TSSs) were annotated and classified with respect to their location relative to coding sequences (CDSs). Predicted TSSs were used to re-annotate 182 CDSs. The TSSs of 2422 CDSs were detected, and consensus promotor logos were derived. Interestingly, many leaderless messenger RNAs (mRNAs) were found. In addition, many mRNAs were transcribed from multiple alternative TSSs. We observed pervasive intragenic TSSs both in sense and antisense to CDSs. Antisense transcripts were enriched near the 5′ end of mRNAs, indicating a functional role in post-transcriptional regulation. In total, 578 TSSs were detected in intergenic regions, of which 35 were identified as putative small regulatory RNAs. Finally, we provide a detailed analysis of the main copper resistance clusters in CH34, which include many intragenic and antisense transcripts. These results clearly highlight the ubiquity of noncoding transcripts in the CH34 transcriptome, many of which are putatively involved in the regulation of metal resistance.

scholarly journals SUT1-promoted sterol uptake involves the ABC transporter Aus1 and the mannoprotein Dan1 whose synergistic action is sufficient for this process

2004 ◽  
Vol 381 (1) ◽  
pp. 195-202 ◽  
Author(s):  
Parissa ALIMARDANI ◽  
Matthieu RÉGNACQ ◽  
Carole MOREAU-VAUZELLE ◽  
Thierry FERREIRA ◽  
Tristan ROSSIGNOL ◽  
...  
Keyword(s):  
Abc Transporter ◽  
Constitutive Expression ◽  
Open Reading Frames ◽  
Wild Type ◽  
Gene Encoding ◽  
Yeast Saccharomyces Cerevisiae ◽  
Sterol Uptake ◽  
Genome Wide ◽  
A Genome

Efficient sterol influx in the yeast Saccharomyces cerevisiae is restricted to anaerobiosis or to haem deficiency resulting from mutations. Constitutive expression of SUT1, an hypoxic gene encoding a transcriptional regulator, induces sterol uptake in aerobiosis. A genome-wide approach using DNA microarray was used to identify the mediators of SUT1 effects on aerobic sterol uptake. A total of 121 ORFs (open reading frames) were significantly and differentially expressed after SUT1 overexpression, 61 down-regulated and 60 up-regulated. Among these genes, the role of the putative ABC transporter (ATP-binding-cassette transporter) Aus1, and of the cell-wall mannoprotein Dan1, was characterized better. These two genes play an essential role in aerobic sterol uptake, since their deletion compromised the SUT1 effects, but individual overexpression of either of these genes in a wild-type background was not sufficient for this process. However, constitutive co-expression of AUS1 and DAN1 in a wild-type background resulted in sterol influx in aerobiosis. These results suggest that the corresponding proteins may act synergistically in vivo to promote sterol uptake.

Heavy metal resistance in Cupriavidus metallidurans CH34 is governed by an intricate transcriptional network

BioMetals ◽  
2011 ◽  
Vol 24 (6) ◽  
pp. 1133-1151 ◽  
Author(s):  
Pieter Monsieurs ◽  
Hugo Moors ◽  
Rob Van Houdt ◽  
Paul J. Janssen ◽  
Ann Janssen ◽  
...  
Keyword(s):  
Heavy Metal ◽  
Heavy Metal Resistance ◽  
Metal Resistance ◽  
Transcriptional Network ◽  
Cupriavidus Metallidurans Ch34 ◽  
Cupriavidus Metallidurans

scholarly journals Cupriavidus metallidurans CH34, a historical perspective on its discovery, characterization and metal resistance

2020 ◽  
Author(s):  
Max Mergeay ◽  
Rob Van Houdt
Keyword(s):  
Type Strain ◽  
Historical Perspective ◽  
Bacterial Resistance ◽  
Metal Resistance ◽  
Early Research ◽  
Molecular Techniques ◽  
Ecological Niches ◽  
Cupriavidus Metallidurans Ch34 ◽  
Resistance Determinants ◽  
Cupriavidus Metallidurans

Abstract Cupriavidus metallidurans, and in particular type strain CH34, became a model bacterium to study bacterial resistance to metals. Although nowadays the routine use of a wide variety of omics and molecular techniques allow refining, deepening and expanding our knowledge on adaptation and resistance to metals, these were not available at the onset of C. metallidurans research starting from its isolation in 1976. This minireview describes the early research and legacy tools used to study its metal resistance determinants, characteristic megaplasmids, ecological niches and environmental applications.

CopK from Cupriavidus metallidurans CH34 Binds Cu(I) in a Tetrathioether Site: Characterization by X-ray Absorption and NMR Spectroscopy

2010 ◽  
Vol 132 (11) ◽  
pp. 3770-3777 ◽  
Author(s):  
Géraldine Sarret ◽  
Adrien Favier ◽  
Jacques Covès ◽  
Jean-Louis Hazemann ◽  
Max Mergeay ◽  
...  
Keyword(s):  
Nmr Spectroscopy ◽  
Site Characterization ◽  
Cupriavidus Metallidurans Ch34 ◽  
X Ray ◽  
Cupriavidus Metallidurans ◽  
X Ray Absorption
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