Loss of Zhf and the tightly regulated zinc-uptake system SpZrt1 inSchizosaccharomyces pombereveals the delicacy of cellular zinc balance

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.

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Functional Analysis of the Bacillus subtilis Zur Regulon

Journal of Bacteriology ◽

10.1128/jb.184.23.6508-6514.2002 ◽

2002 ◽

Vol 184 (23) ◽

pp. 6508-6514 ◽

Cited By ~ 93

Author(s):

Ahmed Gaballa ◽

Tao Wang ◽

Rick W. Ye ◽

John D. Helmann

Keyword(s):

Bacillus Subtilis ◽

Metal Ion ◽

Dna Microarrays ◽

Optimal Growth ◽

Zinc Uptake ◽

Uptake System ◽

Mobility Shift ◽

Transport Pathway ◽

Dnase I Footprinting ◽

Transporter Family

ABSTRACT The Bacillus subtilis zinc uptake repressor (Zur) regulates genes involved in zinc uptake. We have used DNA microarrays to identify genes that are derepressed in a zur mutant. In addition to members of the two previously identified Zur-regulated operons (yciC and ycdHI-yceA), we identified two other genes, yciA and yciB, as targets of Zur regulation. Electrophoretic mobility shift experiments demonstrated that all three operons are direct targets of Zur regulation. Zur binds to an ∼28-bp operator upstream of the yciA gene, as judged by DNase I footprinting, and similar operator sites are found preceding each of the previously described target operons, yciC and ycdHI-yceA. Analysis of a yciA-lacZ fusion indicates that this operon is induced under zinc starvation conditions and derepressed in the zur mutant. Phenotypic analyses suggest that the YciA, YciB, and YciC proteins may function as part of the same Zn(II) transport pathway. Mutation of yciA or yciC, singly or in combination, had little effect on growth of the wild-type strain but significantly impaired the growth of the ycdH mutant under conditions of zinc limitation. Since the YciA, YciB, and YciC proteins are not obviously related to any known transporter family, they may define a new class of metal ion uptake system. Mutant strains lacking all three identified zinc uptake systems (yciABC, ycdHI-yceA, and zosA) are dependent on micromolar levels of added zinc for optimal growth.

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Is Spina Bifida in Man Linked to a Genetic Defect of Cellular Zinc Uptake in Children ?

Trace Elements in Man and Animals 6 ◽

10.1007/978-1-4613-0723-5_219 ◽

1988 ◽

pp. 609-610 ◽

Cited By ~ 1

Author(s):

Alain Favier ◽

Brigitte Dardelet ◽

Marie-Jeanne Richard ◽

Josiane Arnaud

Keyword(s):

Spina Bifida ◽

Genetic Defect ◽

Zinc Uptake ◽

Cellular Zinc

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The zinc transporter ZnuABC is critical for the virulence of Chromobacterium violaceum and contributes to diverse zinc-dependent physiological processes

10.1101/2021.06.03.447022 ◽

2021 ◽

Author(s):

Renato E. R. S. Santos ◽

Waldir P. da Silva Júnior ◽

Simone A. Harrison ◽

Eric P Skaar ◽

Walter J. Chazin ◽

...

Keyword(s):

Mutant Strain ◽

Zinc Uptake ◽

Chromobacterium Violaceum ◽

Host Protein ◽

Infection Model ◽

Uptake System ◽

Environmental Bacterium ◽

Mouse Infection Model ◽

Synthetic Metal

Chromobacterium violaceum is a ubiquitous environmental bacterium that causes sporadic life-threatening infections in humans. How C. violaceum acquires zinc to colonize environmental and host niches is unknown. In this work, we demonstrated that C. violaceum employs the zinc uptake system ZnuABC to overcome zinc limitation in the host, ensuring the zinc supply for several physiological demands. Our data indicated that the C. violaceum ZnuABC transporter is encoded in a zur-CV_RS15045-CV_RS15040-znuCBA operon. This operon was repressed by the zinc uptake regulator Zur and derepressed in the presence of the host protein calprotectin (CP) and the synthetic metal chelator EDTA. A ΔznuCBA mutant strain showed impaired growth under these zinc-chelated conditions. Moreover, the deletion of znuCBA provoked a reduction in violacein production, swimming motility, biofilm formation, and bacterial competition. Remarkably, the ΔznuCBA mutant strain was highly attenuated for virulence in an in vivo mouse infection model and showed a low capacity to colonize the liver, grow in the presence of CP, and resist neutrophil killing. Overall, our findings demonstrate that ZnuABC is essential for C. violaceum virulence, contributing to subvert the zinc-based host nutritional immunity.

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A peroxide-induced zinc uptake system plays an important role in protection against oxidative stress in Bacillus subtilis

Molecular Microbiology ◽

10.1046/j.1365-2958.2002.03068.x ◽

2002 ◽

Vol 45 (4) ◽

pp. 997-1005 ◽

Cited By ~ 95

Author(s):

Ahmed Gaballa ◽

John D. Helmann

Keyword(s):

Oxidative Stress ◽

Bacillus Subtilis ◽

Zinc Uptake ◽

Uptake System

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Coordinated Zinc Homeostasis Is Essential for the Wild-Type Virulence of Brucella abortus

Journal of Bacteriology ◽

10.1128/jb.02543-14 ◽

2015 ◽

Vol 197 (9) ◽

pp. 1582-1591 ◽

Cited By ~ 15

Author(s):

Lauren M. Sheehan ◽

James A. Budnick ◽

R. Martin Roop ◽

Clayton C. Caswell

Keyword(s):

Mouse Model ◽

Brucella Abortus ◽

Zinc Uptake ◽

Zinc Homeostasis ◽

Zinc Toxicity ◽

Uptake System ◽

Bacterial Cells ◽

Gene Encoding ◽

Content Type ◽

Genes Encoding

ABSTRACTMetal homeostasis in bacterial cells is a highly regulated process requiring intricately coordinated import and export, as well as precise sensing of intracellular metal concentrations. The uptake of zinc (Zn) has been linked to the virulence ofBrucella abortus; however, the capacity ofBrucellastrains to sense Zn levels and subsequently coordinate Zn homeostasis has not been described. Here, we show that expression of the genes encoding the zinc uptake system ZnuABC is negatively regulated by the Zn-sensing Fur family transcriptional regulator, Zur, by direct interactions between Zur and the promoter region ofznuABC. Moreover, the MerR-type regulator, ZntR, controls the expression of the gene encoding the Zn exporter ZntA by binding directly to its promoter. Deletion ofzurorzntRalone did not result in increased zinc toxicity in the corresponding mutants; however, deletion ofzntAled to increased sensitivity to Zn but not to other metals, such as Cu and Ni, suggesting that ZntA is a Zn-specific exporter. Strikingly, deletion ofzntRresulted in significant attenuation ofB. abortusin a mouse model of chronic infection, and subsequent experiments revealed that overexpression ofzntAin thezntRmutant is the molecular basis for its decreased virulence.IMPORTANCEThe importance of zinc uptake forBrucellapathogenesis has been demonstrated previously, but to date, there has been no description of how overall zinc homeostasis is maintained and genetically controlled in the brucellae. The present work defines the predominant zinc export system, as well as the key genetic regulators of both zinc uptake and export inBrucella abortus. Moreover, the data show the importance of precise coordination of the zinc homeostasis systems as disregulation of some elements of these systems leads to the attenuation ofBrucellavirulence in a mouse model. Overall, this study advances our understanding of the essential role of zinc in the pathogenesis of intracellular bacteria.

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The zrfA and zrfB Genes of Aspergillus fumigatus Encode the Zinc Transporter Proteins of a Zinc Uptake System Induced in an Acid, Zinc-Depleted Environment

Eukaryotic Cell ◽

10.1128/ec.4.5.837-848.2005 ◽

2005 ◽

Vol 4 (5) ◽

pp. 837-848 ◽

Cited By ~ 55

Author(s):

Rocío Vicentefranqueira ◽

Miguel Ángel Moreno ◽

Fernando Leal ◽

José Antonio Calera

Keyword(s):

Normal Growth ◽

Zinc Uptake ◽

Zinc Transporters ◽

Uptake System ◽

Zinc Transport ◽

Transcriptional Expression ◽

Wild Type ◽

Environmental Concentration ◽

Single Deletion ◽

Immunodominant Antigens

ABSTRACT Zinc is an essential micronutrient that cells must obtain from the environment in order to develop their normal growth. Previous work performed at our laboratory showed that the synthesis of immunodominant antigens from Aspergillus spp., including A. fumigatus, was up-regulated by a low environmental concentration of zinc. These results suggested that a tightly regulated system for the fungus to grow under zinc-limiting conditions must underlie the ability of A. fumigatus to acquire zinc in such environments. In this work, we show that zrfA and zrfB are two of the genes that encode membrane zinc transporters from A. fumigatus in this system. Expression of these genes is differentially down-regulated by increasing concentrations of zinc in the medium. Thus, the transcription of zrfB is turned off at a concentration 50-fold higher than that for zrfA transcription. In addition, phenotypic analyses of single zrfAΔ and zrfBΔ mutants and a double zrfAzrfBΔ mutant revealed that the deletion of zrfB causes a greater defect in growth than the single deletion of zrfA. Deletion of both genes has a dramatic effect on growth under acid, zinc-limiting conditions. Interestingly, in neutral or slightly alkaline zinc-depleted medium, the transcriptional expression of both genes is down-regulated to such an extent that even in the absence of a supplement of zinc, the expression of zrfA and zrfB is strongly reduced. This fact correlates with the growth observed in alkaline medium, in which even a zrfAzrfBΔ double mutant was able to grow in a similar way to the wild-type under extremely zinc-limiting conditions. In sum, the zinc transport proteins encoded by zrfA and zrfB are members of a zinc uptake system of A. fumigatus that operates mainly under acid, zinc-limiting conditions.

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