scholarly journals Identification and functional characterization of a novel Candida albicans gene CaMNN5 that suppresses the iron-dependent growth defect of Saccharomyces cerevisiae aft1Δ mutant

2005 ◽  
Vol 389 (1) ◽  
pp. 27-35 ◽  
Author(s):  
Chen BAI ◽  
Fong Yee CHAN ◽  
Yue WANG
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Candida Albicans ◽  
Iron Uptake ◽  
Iron Transport ◽  
Lucifer Yellow ◽  
Functional Characterization ◽  
Endocytic Pathway ◽  
Transport Systems ◽  
Growth Defect ◽  
Iron Starvation

In Saccharomyces cerevisiae, the transcription factor Aft1p plays a central role in regulating many genes involved in iron acquisition and utilization. An aft1Δ mutant exhibits severely retarded growth under iron starvation. To identify the functional counterpart of AFT1 in Candida albicans, we transformed a C. albicans genomic DNA library into aft1Δ to isolate genes that could allow the mutant to grow under iron-limiting conditions. In the present paper, we describe the unexpected discovery in this screen of CaMNN5. CaMnn5p is an α-1,2-mannosyltransferease, but its growth-promoting function in iron-limiting conditions does not require this enzymatic activity. Its function is also independent of the high-affinity iron transport systems that are mediated by Ftr1p and Fth1p. We obtained evidence suggesting that CaMnn5p may function along the endocytic pathway, because it cannot promote the growth of end4Δ and vps4Δ mutants, where the endocytic pathway is blocked at an early and late step respectively. Neither can it promote the growth of a fth1Δ smf3Δ mutant, where the vacuole–cytosol iron transport is blocked. Expression of CaMNN5 in S. cerevisiae specifically enhances an endocytosis-dependent mechanism of iron uptake without increasing the uptake of Lucifer Yellow, a marker for fluid-phase endocytosis. CaMnn5p contains three putative Lys-Glu-Xaa-Xaa-Glu iron-binding sites and co-immunoprecipitates with 55Fe. We propose that CaMnn5p promotes iron uptake and usage along the endocytosis pathway under iron-limiting conditions, a novel function that might have evolved in C. albicans.

scholarly journals Genes Essential to Iron Transport in the Cyanobacterium Synechocystis sp. Strain PCC 6803

2001 ◽  
Vol 183 (9) ◽  
pp. 2779-2784 ◽  
Author(s):  
Hirokazu Katoh ◽  
Natsu Hagino ◽  
Arthur R. Grossman ◽  
Teruo Ogawa
Keyword(s):  
Iron Uptake ◽  
Iron Transport ◽  
Ferric Iron ◽  
Iron Acquisition ◽  
Complete Medium ◽  
Iron Starvation ◽  
Transporter Family ◽  
Genes Encoding ◽  
In Cells ◽  
Pcc 6803

ABSTRACT Genes encoding polypeptides of an ATP binding cassette (ABC)-type ferric iron transporter that plays a major role in iron acquisition inSynechocystis sp. strain PCC 6803 were identified. These genes are slr1295, slr0513, slr0327, and recently reportedsll1878 (Katoh et al., J. Bacteriol. 182:6523–6524, 2000) and were designated futA1, futA2, futB, andfutC, respectively, for their involvement in ferric iron uptake. Inactivation of these genes individually or futA1and futA2 together greatly reduced the activity of ferric iron uptake in cells grown in complete medium or iron-deprived medium. All the fut genes are expressed in cells grown in complete medium, and expression was enhanced by iron starvation. ThefutA1 and futA2 genes appear to encode periplasmic proteins that play a redundant role in iron binding. The deduced products of futB and futC genes contain nucleotide-binding motifs and belong to the ABC transporter family of inner-membrane-bound and membrane-associated proteins, respectively. These results and sequence similarities among the four genes suggest that the Fut system is related to the Sfu/Fbp family of iron transporters. Inactivation of slr1392, a homologue offeoB in Escherichia coli, greatly reduced the activity of ferrous iron transport. This system is induced by intracellular low iron concentrations that are achieved in cells exposed to iron-free medium or in the fut-less mutants grown in complete medium.

scholarly journals Characterization of iron-binding motifs in Candida albicans high-affinity iron permease CaFtr1p by site-directed mutagenesis

2002 ◽  
Vol 368 (2) ◽  
pp. 641-647 ◽  
Author(s):  
Hao-Ming FANG ◽  
Yue WANG
Keyword(s):  
Amino Acids ◽  
Candida Albicans ◽  
Iron Uptake ◽  
Iron Transport ◽  
Site Directed Mutagenesis ◽  
Detectable Effect ◽  
Directed Mutagenesis ◽  
Binding Motif ◽  
Iron Binding ◽  
Direct Binding

A peptide motif Glu-Xaa-Xaa-Glu has been implicated in direct binding of ferric iron in several proteins involved in iron transport, sensing or storage. However, it is not known whether the motif alone is sufficient for iron binding and whether functional replacement of the conserved residues by other amino acids with similar properties is possible. We previously identified a Candida albicans iron permease, CaFtr1p, which contains five Glu-Xaa-Xaa-Glu motifs [Ramanan and Wang (2000) Science 288, 1062—1065]. In this study, we investigated the role of each of these motifs in iron uptake by site-directed mutagenesis. Substitution of Ala for any one of the two Glu residues in Glu-Gly-Leu-Glu158—161 abolished iron-uptake activity, while the same substitution in any of the other four motifs had little effect, indicating that only the motif at position 158—161 is required for iron transport. We then evaluated the importance of each of the residues within and immediately adjacent to this motif in iron uptake. The permease remained active when any one of the Glu residues was replaced by Asp, while it became inactive when both were replaced. We also found that the amino acid immediately in front of Glu-Gly-Leu-Glu158—161 must be either Arg or Lys. In addition, substitution of any of the two residues in the middle with several structurally distinct amino acids had no detectable effect on iron uptake. Here we propose to extend the iron-binding motif to Arg/Lys-Glu/Asp-Xaa-Xaa-Glu or Arg/Lys-Glu-Xaa-Xaa-Glu/Asp, which may serve as a guide for the identification of potential iron-binding sites in proteins.

scholarly journals Hierarchy of Iron Uptake Systems: Yfu and Yiu Are Functional in Yersinia pestis

2006 ◽  
Vol 74 (11) ◽  
pp. 6171-6178 ◽  
Author(s):  
Olga Kirillina ◽  
Alexander G. Bobrov ◽  
Jacqueline D. Fetherston ◽  
Robert D. Perry
Keyword(s):  
Yersinia Pestis ◽  
Iron Uptake ◽  
Disease Process ◽  
Transport Systems ◽  
Growth Defect ◽  
Uptake System ◽  
Outer Membrane Receptor ◽  
Abc Transport ◽  
Iron Deficient ◽  
Additional Iron

ABSTRACT In addition to the yersiniabactin (Ybt) siderophore-dependent system, two inorganic iron ABC transport systems of Yersinia pestis, Yfe and Yfu, have been characterized. Here we show that the Yfu system functions in Y. pestis: a Ybt− Yfe− Yfu− mutant exhibited a greater growth defect under iron-deficient conditions than its Ybt− Yfe− parental strain. We also demonstrate that another putative Y. pestis iron uptake system, Yiu, which potentially encodes an outer membrane receptor, YiuR, and an ABC iron transport cassette, YiuABC, is functional. The cloned yiuABC operon restored growth of an enterobactin-deficient mutant Escherichia coli strain, 1017, under iron-chelated conditions. Iron uptake by the Yiu system in Y. pestis was demonstrated only when the Ybt, Yfe, and Yfu systems were mutated. Using a yiuA::lacZ fusion, we show that the yiuABC promoter is repressed by iron through Fur. A mouse model of bubonic plague failed to show a significant role for the Yiu system in the disease process. These results demonstrate that two additional iron transporters are functional in Y. pestis and indicate that there is a hierarchy of iron transporters, with Ybt being most effective and Yiu being the least effective of those systems which have been characterized.

scholarly journals The F-Box Protein Rcy1p Is Involved in Endocytic Membrane Traffic and Recycling Out of an Early Endosome in Saccharomyces cerevisiae

2000 ◽  
Vol 149 (2) ◽  
pp. 397-410 ◽  
Author(s):  
Andreas Wiederkehr ◽  
Sandrine Avaro ◽  
Cristina Prescianotto-Baschong ◽  
Rosine Haguenauer-Tsapis ◽  
Howard Riezman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Membrane Trafficking ◽  
Fluorescent Dyes ◽  
Lucifer Yellow ◽  
Endocytic Pathway ◽  
Major Fraction ◽  
Membrane Bound ◽  
Recycling Pathway ◽  
F Box Protein ◽  
Vacuolar Protein

In Saccharomyces cerevisiae, endocytic material is transported through different membrane-bound compartments before it reaches the vacuole. In a screen for mutants that affect membrane trafficking along the endocytic pathway, we have identified a novel mutant disrupted for the gene YJL204c that we have renamed RCY1 (recycling 1). Deletion of RCY1 leads to an early block in the endocytic pathway before the intersection with the vacuolar protein sorting pathway. Mutation of RCY1 leads to the accumulation of an enlarged compartment that contains the t-SNARE Tlg1p and lies close to areas of cell expansion. In addition, endocytic markers such as Ste2p and the fluorescent dyes, Lucifer yellow and FM4-64, were found in a similar enlarged compartment after their internalization. To determine whether rcy1Δ is defective for recycling, we have developed an assay that measures the recycling of previously internalized FM4-64. This method enables us to follow the recycling pathway in yeast in real time. Using this assay, it could be demonstrated that recycling of membranes is rapid in S. cerevisiae and that a major fraction of internalized FM4-64 is secreted back into the medium within a few minutes. The rcy1Δ mutant is strongly defective in recycling.

scholarly journals Functional characterization of the ferroxidase, permease high-affinity iron transport complex from Candida albicans

2011 ◽  
Vol 81 (2) ◽  
pp. 473-485 ◽  
Author(s):  
Lynn Ziegler ◽  
Alaina Terzulli ◽  
Ruchi Gaur ◽  
Ryan McCarthy ◽  
Daniel J. Kosman
Keyword(s):  
Candida Albicans ◽  
Iron Transport ◽  
Functional Characterization ◽  
High Affinity ◽  
Transport Complex

scholarly journals Iron Transport Systems in Neisseria meningitidis

2004 ◽  
Vol 68 (1) ◽  
pp. 154-171 ◽  
Author(s):  
Donna Perkins-Balding ◽  
Melanie Ratliff-Griffin ◽  
Igor Stojiljkovic
Keyword(s):  
Neisseria Meningitidis ◽  
Iron Uptake ◽  
Iron Transport ◽  
Current Understanding ◽  
Transport Systems ◽  
Transport Activity ◽  
Host Proteins ◽  
Human Host ◽  
High Affinity ◽  
Dependent Transport

SUMMARY Acquisition of iron and iron complexes has long been recognized as a major determinant in the pathogenesis of Neisseria meningitidis. In this review, high-affinity iron uptake systems, which allow meningococci to utilize the human host proteins transferrin, lactoferrin, hemoglobin, and haptoglobin-hemoglobin as sources of essential iron, are described. Classic features of bacterial iron transport systems, such as regulation by the iron-responsive repressor Fur and TonB-dependent transport activity, are discussed, as well as more specific features of meningococcal iron transport. Our current understanding of how N. meningitidis acquires iron from the human host and the vaccine potentials of various components of these iron transport systems are also reviewed.

scholarly journals Genetic evidence that ferric reductase is required for iron uptake in Saccharomyces cerevisiae.

1990 ◽  
Vol 10 (5) ◽  
pp. 2294-2301 ◽  
Author(s):  
A Dancis ◽  
R D Klausner ◽  
A G Hinnebusch ◽  
J G Barriocanal
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Plasma Membrane ◽  
Iron Uptake ◽  
Reductase Activity ◽  
Gene Transcript ◽  
Single Mutation ◽  
Uptake System ◽  
Ferric Reductase ◽  
Wild Type ◽  
Iron Starvation

The requirement for a reduction step in cellular iron uptake has been postulated, and the existence of plasma membrane ferric reductase activity has been described in both procaryotic and eucaryotic cells. In the yeast Saccharomyces cerevisiae, there is an externally directed reductase activity that is regulated by the concentration of iron in the growth medium; maximal activity is induced by iron starvation. We report here the isolation of a mutant of S. cerevisiae lacking the reductase activity. This mutant is deficient in the uptake of ferric iron and is extremely sensitive to iron deprivation. Genetic analysis of the mutant demonstrates that the reductase and ferric uptake deficiencies are due to a single mutation that we designate fre1-1. Both phenotypes cosegregate in meiosis, corevert with a frequency of 10(-7), and are complemented by a 3.5-kilobase fragment of genomic DNA from wild-type S. cerevisiae. This fragment contains FRE1, the wild-type allele of the mutant gene. The level of the gene transcript is regulated by iron in the same was as the reductase activity. The ferrous ion product of the reductase must traverse the plasma membrane. A high-affinity (Km = 5 microM) ferrous uptake system is present in both wild-type and mutant cells. Thus, iron uptake in S. cerevisiae is mediated by two plasma membrane components, a reductase and a ferrous transport system.

scholarly journals Overlap of copper and iron uptake systems in mitochondria in Saccharomyces cerevisiae

Open Biology ◽  
2016 ◽  
Vol 6 (1) ◽  
pp. 150223 ◽  
Author(s):  
Katherine E. Vest ◽  
Jing Wang ◽  
Micah G. Gammon ◽  
Margaret K. Maynard ◽  
Olivia L. White ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transport Systems ◽  
Growth Defect ◽  
Intermembrane Space ◽  
Copper Uptake ◽  
Copper Sensor ◽  
The Matrix ◽  
Mitochondrial Carrier Family ◽  
Dependent Growth ◽  
Copper Import

In Saccharomyces cerevisiae , the mitochondrial carrier family protein Pic2 imports copper into the matrix. Deletion of PIC2 causes defects in mitochondrial copper uptake and copper-dependent growth phenotypes owing to decreased cytochrome c oxidase activity. However, copper import is not completely eliminated in this mutant, so alternative transport systems must exist. Deletion of MRS3 , a component of the iron import machinery, also causes a copper-dependent growth defect on non-fermentable carbon. Deletion of both PIC2 and MRS3 led to a more severe respiratory growth defect than either individual mutant. In addition, MRS3 expressed from a high copy number vector was able to suppress the oxygen consumption and copper uptake defects of a strain lacking PIC2 . When expressed in Lactococcus lactis , Mrs3 mediated copper and iron import. Finally, a PIC2 and MRS3 double mutant prevented the copper-dependent activation of a heterologously expressed copper sensor in the mitochondrial intermembrane space. Taken together, these data support a role for the iron transporter Mrs3 in copper import into the mitochondrial matrix.

scholarly journals Spt3 Plays Opposite Roles in Filamentous Growth in Saccharomyces cerevisiae and Candida albicans and Is Required for C. albicans Virulence

Genetics ◽  
2002 ◽  
Vol 161 (2) ◽  
pp. 509-519
Author(s):  
Lisa Laprade ◽  
Victor L Boyartchuk ◽  
William F Dietrich ◽  
Fred Winston
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Candida Albicans ◽  
Filamentous Growth ◽  
Growth Defect ◽  
Invasive Growth ◽  
Pathogenic Yeast ◽  
Normal Expression

Abstract Spt3 of Saccharomyces cerevisiae is required for the normal transcription of many genes in vivo. Past studies have shown that Spt3 is required for both mating and sporulation, two events that initiate when cells are at G1/START. We now show that Spt3 is needed for two other events that begin at G1/START, diploid filamentous growth and haploid invasive growth. In addition, Spt3 is required for normal expression of FLO11, a gene required for filamentous growth, although this defect is not the sole cause of the spt3Δ/spt3Δ filamentous growth defect. To extend our studies of Spt3's role in filamentous growth to the pathogenic yeast Candida albicans, we have identified the C. albicans SPT3 gene and have studied its role in C. albicans filamentous growth and virulence. Surprisingly, C. albicans spt3Δ/spt3Δ mutants are hyperfilamentous, the opposite phenotype observed for S. cerevisiae spt3Δ/spt3Δ mutants. Furthermore, C. albicans spt3Δ/spt3Δ mutants are avirulent in mice. These experiments demonstrate that Spt3 plays important but opposite roles in filamentous growth in S. cerevisiae and C. albicans.

scholarly journals Disruption of a Novel Iron Transport System Reverses Oxidative Stress Phenotypes of a dpr Mutant Strain of Streptococcus mutans

2018 ◽  
Vol 200 (14) ◽  
Author(s):  
Tridib Ganguly ◽  
Jessica K. Kajfasz ◽  
James H. Miller ◽  
Eric Rabinowitz ◽  
Lívia C. C. Galvão ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Streptococcus Mutans ◽  
Hydroxyl Radicals ◽  
Transport System ◽  
Iron Uptake ◽  
Iron Transport ◽  
Transport Systems ◽  
Phenotypic Characterization ◽  
Content Type

ABSTRACT The Dps-like peroxide resistance protein (Dpr) is essential for H 2 O 2 stress tolerance and aerobic growth of the oral pathogen Streptococcus mutans . Dpr accumulates during oxidative stress, protecting the cell by sequestering iron ions and thereby preventing the generation of toxic hydroxyl radicals that result from the interaction of iron with H 2 O 2 . Previously, we reported that the SpxA1 and SpxA2 regulators positively regulate expression of dpr in S. mutans . Using an antibody raised against S. mutans Dpr, we confirmed at the protein level the central and cooperative nature of SpxA1 and SpxA2 regulation in Dpr production. During phenotypic characterization of the S. mutans Δ dpr strain, we observed the appearance of distinct colony variants, which sometimes lost the oxidative stress sensitivity typical of Δ dpr strains. Whole-genome sequencing of these phenotypically distinct Δ dpr isolates revealed that a putative iron transporter operon, smu995-smu998 , was a genomic hot spot with multiple single nucleotide polymorphisms identified within the different isolates. Deletion of smu995 or the entire smu995-smu998 operon in the Δ dpr background strain completely reversed the oxidative stress-sensitive phenotypes associated with dpr inactivation. Conversely, inactivation of genes encoding the ferrous iron transport system FeoABC did not alleviate phenotypes of the Δ dpr strain. Preliminary characterization of strains lacking smu995-smu998 , feoABC , and the iron/manganese transporter gene sloABC revealed the interactive nature of these three systems in iron transport but also indicated that there may be additional iron uptake systems in S. mutans . IMPORTANCE The dental caries-associated pathogen Streptococcus mutans routinely encounters oxidative stress within the human plaque biofilm. Previous studies revealed that the iron-binding protein Dpr confers protection toward oxidative stress by limiting free iron availability, which is associated with the generation of toxic hydroxyl radicals. Here, we report the identification of spontaneously occurring mutations within Δ dpr strains. Several of those mutations were mapped to the operon smu995-smu998 , revealing a previously uncharacterized system that appears to be important in iron acquisition. Disruption of the smu995-smu998 operon resulted in reversion of the stress-sensitive phenotype typical of a Δ dpr strain. Our data suggest that the Smu995-Smu998 system works along with other known metal transport systems of S. mutans , i.e., FeoABC and SloABC, to coordinate iron uptake.

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