scholarly journals Multiple Evolutionarily Conserved Domains of Cap2 Are Required for Promoter Recruitment and Iron Homeostasis Gene Regulation

mSphere ◽  
2018 ◽  
Vol 3 (4) ◽  
Author(s):  
Manjit Kumar Srivastav ◽  
Neha Agarwal ◽  
Krishnamurthy Natarajan
Keyword(s):  
Iron Uptake ◽  
Transcriptional Activity ◽  
Iron Homeostasis ◽  
Protein Domains ◽  
Conserved Domains ◽  
Content Type ◽  
Human Fungal Pathogen ◽  
Bzip Domain ◽  
Evolutionarily Conserved ◽  
Promoter Recruitment

ABSTRACTIron is required for growth and metabolism by virtually all organisms. The human fungal pathogenCandida albicanshas evolved multiple strategies to acquire iron. The Cap2/Hap43 transcriptional regulator, essential for robust virulence ofC. albicans, controls iron homeostasis gene expression by promoter binding and repression of iron utilization genes. The expression of iron uptake genes is also dependent on Cap2, although Cap2 was not recruited to its promoters. Cap2, bearing the conserved bipartite HAP4L-bZIP domain, also contains multiple blocks of amino acids that form the highly conserved carboxyl-terminal region. In this study, we sought to identify the requirements of the different domains for Cap2 function. We constructed a series of mutants bearing either point mutations or deletions in the conserved domains and examined Cap2 activity. Deletion of the highly conserved extreme C-terminal region did not impair expression of Cap2 mutant protein but impaired cell growth and expression of iron homeostasis genes under iron-depleted conditions. Mutations in the amino-terminal HAP4L and basic leucine zipper (bZIP) domains also impaired growth and gene expression. Furthermore, chromatin immunoprecipitation (ChIP) assays showed that the HAP4L domain and the bZIP domain are both essential for Cap2 recruitment toACO1andCYC1promoters. Unexpectedly, the C-terminal conserved region was also essential for Cap2 promoter recruitment. Thus, our results suggest that Cap2 employs multiple evolutionarily conserved domains, including the C-terminal domain for its transcriptional activity.IMPORTANCEIron is an essential micronutrient for living cells.Candida albicans, the predominant human fungal pathogen, thrives under diverse environments with vastly different iron levels in the mammalian host. Therefore, to tightly control iron homeostasis,C. albicanshas evolved a set of transcriptional regulators that cooperate to either upregulate or downregulate transcription of iron uptake genes or iron utilization genes. Cap2/Hap43, a critical transcriptional regulator, contains multiple conserved protein domains. In this study, we carried out mutational analyses to identify the functional roles of the conserved protein domains in Cap2. Our results show that the bZIP, HAP4L, and the C-terminal domain are each required for Cap2 transcriptional activity. Thus, Cap2 employs multiple, disparate protein domains for regulation of iron homeostasis inC. albicans.

scholarly journals Complex Iron Uptake by the Putrebactin-Mediated and Feo Systems in Shewanella oneidensis

2018 ◽  
Vol 84 (20) ◽  
Author(s):  
Lulu Liu ◽  
Shisheng Li ◽  
Sijing Wang ◽  
Ziyang Dong ◽  
Haichun Gao
Keyword(s):  
Iron Uptake ◽  
Iron Homeostasis ◽  
Shewanella Oneidensis ◽  
Special Role ◽  
Uptake System ◽  
Biological Processes ◽  
Content Type ◽  
Iron Uptake System ◽  
Reducing Bacteria ◽  
Metal Reducing Bacteria

ABSTRACT Shewanella oneidensis is an extensively studied bacterium capable of respiring minerals, including a variety of iron ores, as terminal electron acceptors (EAs). Although iron plays an essential and special role in iron respiration of S. oneidensis, little has been done to date to investigate the characteristics of iron transport in this bacterium. In this study, we found that all proteins encoded by the pub-putA-putB cluster for putrebactin (S. oneidensis native siderophore) synthesis (PubABC), recognition-transport of Fe3+-putrebactin across the outer membrane (PutA), and reduction of ferric putrebactin (PutB) are essential to putrebactin-mediated iron uptake. Although homologs of PutA are many, none can function as its replacement, but some are able to work with other bacterial siderophores. We then showed that Fe2+-specific Feo is the other primary iron uptake system, based on the synthetical lethal phenotype resulting from the loss of both iron uptake routes. The role of the Feo system in iron uptake appears to be more critical, as growth is significantly impaired by the absence of the system but not of putrebactin. Furthermore, we demonstrate that hydroxyl acids, especially α-types such as lactate, promote iron uptake in a Feo-dependent manner. Overall, our findings underscore the importance of the ferrous iron uptake system in metal-reducing bacteria, providing an insight into iron homeostasis by linking these two biological processes. IMPORTANCE S. oneidensis is among the first- and the best-studied metal-reducing bacteria, with great potential in bioremediation and biotechnology. However, many questions regarding mechanisms closely associated with those applications, such as iron homeostasis, including iron uptake, export, and regulation, remain to be addressed. Here we show that Feo is a primary player in iron uptake in addition to the siderophore-dependent route. The investigation also resolved a few puzzles regarding the unexpected phenotypes of the putA mutant and lactate-dependent iron uptake. By elucidating the physiological roles of these two important iron uptake systems, this work revealed the breadth of the impacts of iron uptake systems on the biological processes.

scholarly journals Dopamine Is a Siderophore-Like Iron Chelator That PromotesSalmonella entericaSerovar Typhimurium Virulence in Mice

mBio ◽  
2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Stefanie Dichtl ◽  
Egon Demetz ◽  
David Haschka ◽  
Piotr Tymoszuk ◽  
Verena Petzer ◽  
...  
Keyword(s):  
Bacterial Growth ◽  
Iron Uptake ◽  
Iron Homeostasis ◽  
Iron Acquisition ◽  
Lipocalin 2 ◽  
Intracellular Iron ◽  
Content Type ◽  
Hepcidin Expression

ABSTRACTWe have recently shown that the catecholamine dopamine regulates cellular iron homeostasis in macrophages. As iron is an essential nutrient for microbes, and intracellular iron availability affects the growth of intracellular bacteria, we studied whether dopamine administration impacts the course ofSalmonellainfections. Dopamine was found to promote the growth ofSalmonellaboth in culture and within bone marrow-derived macrophages, which was dependent on increased bacterial iron acquisition. Dopamine administration to mice infected withSalmonella entericaserovar Typhimurium resulted in significantly increased bacterial burdens in liver and spleen, as well as reduced survival. The promotion of bacterial growth by dopamine was independent of the siderophore-binding host peptide lipocalin-2. Rather, dopamine enhancement of iron uptake requires both the histidine sensor kinase QseC and bacterial iron transporters, in particular SitABCD, and may also involve the increased expression of bacterial iron uptake genes. Deletion or pharmacological blockade of QseC reduced but did not abolish the growth-promoting effects of dopamine. Dopamine also modulated systemic iron homeostasis by increasing hepcidin expression and depleting macrophages of the iron exporter ferroportin, which enhanced intracellular bacterial growth.Salmonellalacking all central iron uptake pathways failed to benefit from dopamine treatment. These observations are potentially relevant to critically ill patients, in whom the pharmacological administration of catecholamines to improve circulatory performance may exacerbate the course of infection with siderophilic bacteria.IMPORTANCEHere we show that dopamine increases bacterial iron incorporation and promotesSalmonellaTyphimurium growth bothin vitroandin vivo. These observations suggest the potential hazards of pharmacological catecholamine administration in patients with bacterial sepsis but also suggest that the inhibition of bacterial iron acquisition might provide a useful approach to antimicrobial therapy.

scholarly journals Iron Homeostasis in Mycobacterium tuberculosis: Mechanistic Insights into Siderophore-Mediated Iron Uptake

2016 ◽  
Vol 198 (18) ◽  
pp. 2399-2409 ◽  
Author(s):  
Manjula Sritharan
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Iron Uptake ◽  
Metal Ion ◽  
Iron Homeostasis ◽  
Iron Status ◽  
Mammalian Host ◽  
Intracellular Iron ◽  
Comprehensive Overview ◽  
Content Type ◽  
Excess Iron

Mycobacterium tuberculosisrequires iron for normal growth but faces a limitation of the metal ion due to its low solubility at biological pH and the withholding of iron by the mammalian host. The pathogen expresses the Fe3+-specific siderophores mycobactin and carboxymycobactin to chelate the metal ion from insoluble iron and the host proteins transferrin, lactoferrin, and ferritin. Siderophore-mediated iron uptake is essential for the survival ofM. tuberculosis, as knockout mutants, which were defective in siderophore synthesis or uptake, failed to survive in low-iron medium and inside macrophages. But as excess iron is toxic due to its catalytic role in the generation of free radicals, regulation of iron uptake is necessary to maintain optimal levels of intracellular iron. The focus of this review is to present a comprehensive overview of iron homeostasis inM. tuberculosisthat is discussed in the context of mycobactin biosynthesis, transport of iron across the mycobacterial cell envelope, and storage of excess iron. The clinical significance of the serum iron status and the expression of the iron-regulated protein HupB in tuberculosis (TB) patients is presented here, highlighting the potential of HupB as a marker, notably in extrapulmonary TB cases.

scholarly journals Comparative Evolution of Morphological Regulatory Functions in Candida Species

2013 ◽  
Vol 12 (10) ◽  
pp. 1356-1368 ◽  
Author(s):  
Erika Lackey ◽  
Geethanjali Vipulanandan ◽  
Delma S. Childers ◽  
David Kadosh
Keyword(s):  
Target Genes ◽  
Pathogenic Fungi ◽  
Specific Gene ◽  
Content Type ◽  
Environmental Signals ◽  
Human Fungal Pathogen ◽  
Evolutionarily Conserved ◽  
Morphological Transitions ◽  
High Level ◽  
Regulatory Functions

ABSTRACTMorphological transitions play an important role in virulence and virulence-related processes in a wide variety of pathogenic fungi, including the most commonly isolated human fungal pathogenCandida albicans. While environmental signals, transcriptional regulators, and target genes associated withC. albicansmorphogenesis are well-characterized, considerably little is known about morphological regulatory mechanisms and the extent to which they are evolutionarily conserved in less pathogenic and less filamentous non-albicans Candidaspecies (NACS). We have identified specific optimal filament-inducing conditions for three NACS (C. tropicalis,C. parapsilosis, andC. guilliermondii), which are very limited, suggesting that these species may be adapted for niche-specific filamentation in the host. Only a subset of evolutionarily conservedC. albicansfilament-specific target genes were induced upon filamentation inC. tropicalis,C. parapsilosis, andC. guilliermondii. One of the genes showing conserved expression wasUME6, a key filament-specific regulator ofC. albicanshyphal development. Constitutive high-level expression ofUME6was sufficient to drive increased filamentation as well as biofilm formation and partly restore conserved filament-specific gene expression in bothC. tropicalisandC. parapsilosis, suggesting that evolutionary differences in filamentation ability among pathogenicCandidaspecies may be partially attributed to alterations in the expression level of a conserved filamentous growth machinery. In contrast toUME6,NRG1, an important repressor ofC. albicansfilamentation, showed only a partly conserved role in controlling NACS filamentation. Overall, our results suggest thatC. albicansmorphological regulatory functions are partially conserved in NACS and have evolved to respond to more specific sets of host environmental cues.

scholarly journals Impact of Na+-Translocating NADH:Quinone Oxidoreductase on Iron Uptake and nqrM Expression in Vibrio cholerae

2019 ◽  
Vol 202 (3) ◽  
Author(s):  
Shubhangi Agarwal ◽  
Melanie Bernt ◽  
Charlotte Toulouse ◽  
Hannes Kurz ◽  
Jens Pfannstiel ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Iron Uptake ◽  
Iron Homeostasis ◽  
Mrna Level ◽  
Strong Impact ◽  
Substrate Uptake ◽  
Uptake System ◽  
Content Type ◽  
Respiratory Enzyme ◽  
Nadh:Quinone Oxidoreductase

ABSTRACT The Na+ ion-translocating NADH:quinone oxidoreductase (NQR) from Vibrio cholerae is a membrane-bound respiratory enzyme which harbors flavins and Fe-S clusters as redox centers. The NQR is the main producer of the sodium motive force (SMF) and drives energy-dissipating processes such as flagellar rotation, substrate uptake, ATP synthesis, and cation-proton antiport. The NQR requires for its maturation, in addition to the six structural genes nqrABCDEF, a flavin attachment gene, apbE, and the nqrM gene, presumably encoding a Fe delivery protein. We here describe growth studies and quantitative real-time PCR for the V. cholerae O395N1 wild-type (wt) strain and its mutant Δnqr and ΔubiC strains, impaired in respiration. In a comparative proteome analysis, FeoB, the membrane subunit of the uptake system for Fe2+ (Feo), was increased in V. cholerae Δnqr. In this study, the upregulation was confirmed on the mRNA level and resulted in improved growth rates of V. cholerae Δnqr with Fe2+ as an iron source. We studied the expression of feoB on other respiratory enzyme deletion mutants such as the ΔubiC mutant to determine whether iron transport is specific to the absence of NQR resulting from impaired respiration. We show that the nqr operon comprises, in addition to the structural nqrABCDEF genes, the downstream apbE and nqrM genes on the same operon and demonstrate induction of the nqr operon by iron in V. cholerae wt. In contrast, expression of the nqrM gene in V. cholerae Δnqr is repressed by iron. The lack of functional NQR has a strong impact on iron homeostasis in V. cholerae and demonstrates that central respiratory metabolism is interwoven with iron uptake and regulation. IMPORTANCE Investigating strategies of iron acquisition, storage, and delivery in Vibrio cholerae is a prerequisite to understand how this pathogen thrives in hostile, iron-limited environments such as the human host. In addition to highlighting the maturation of the respiratory complex NQR, this study points out the influence of NQR on iron metabolism, thereby making it a potential drug target for antibiotics.

scholarly journals Fungicidal Monoclonal Antibody C7 Interferes with Iron Acquisition in Candida albicans

2011 ◽  
Vol 55 (7) ◽  
pp. 3156-3163 ◽  
Author(s):  
Sonia Brena ◽  
Jonathan Cabezas-Olcoz ◽  
María D. Moragues ◽  
Iñigo Fernández de Larrinoa ◽  
Angel Domínguez ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Candida Albicans ◽  
Iron Uptake ◽  
Iron Homeostasis ◽  
Iron Acquisition ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Content Type ◽  
Related Group ◽  
Upregulated Genes

ABSTRACTWe have developed a monoclonal antibody (MAb), C7, that reacts with the Als3p and enolase present in theCandida albicanscell wall and exerts three anti-Candidaactivities: candidacidal activity and inhibition of both adhesion and filamentation. To investigate the mode of action of MAb C7 on fungal viability, we examined changes in the genome-wide gene expression profile ofC. albicansgrown in the presence of a subinhibitory concentration of MAb C7 (12.5 μg/ml) by using microarrays. A total of 49 genes were found to be differentially expressed upon treatment with MAb C7. Of these, 28 were found to be upregulated and 21 were found to be downregulated. The categories of upregulated genes with the largest number of variations were those involved in iron uptake or related to iron homeostasis (42.86%), while the energy-related group accounted for 38.10% of the downregulated genes (8/21). Results were validated by real-time PCR. Since these effects resembled those found under iron-limited conditions, the activity of MAb C7 onC. albicansmutants with deletions in key genes implicated in the three iron acquisition systems described in this yeast was also assessed. Only mutants lacking theTPK1gene and, to a lesser extent, theTPK2gene were less sensitive to the candidacidal effect of MAb C7. FeCl3or hemin at concentrations of ≥7.8 μM reversed the candidacidal effect of MAb C7 onC. albicansin a concentration-dependent manner. The results presented in this study provide evidence that the candidacidal effect of MAb C7 is related to the blockage of the reductive iron uptake pathway ofC. albicans.

scholarly journals Tailoring a Global Iron Regulon to a Uropathogen

mBio ◽  
2020 ◽  
Vol 11 (2) ◽  
Author(s):  
Rajdeep Banerjee ◽  
Erin Weisenhorn ◽  
Kevin J. Schwartz ◽  
Kevin S. Myers ◽  
Jeremy D. Glasner ◽  
...  
Keyword(s):  
Amino Acids ◽  
Urinary Tract ◽  
Amino Acid ◽  
Regulatory Network ◽  
Iron Homeostasis ◽  
Iron Limitation ◽  
Content Type ◽  
E Coli ◽  
General Stress ◽  
K 12

ABSTRACT Pathogenicity islands and plasmids bear genes for pathogenesis of various Escherichia coli pathotypes. Although there is a basic understanding of the contribution of these virulence factors to disease, less is known about variation in regulatory networks in determining disease phenotypes. Here, we dissected a regulatory network directed by the conserved iron homeostasis regulator, ferric uptake regulator (Fur), in uropathogenic E. coli (UPEC) strain CFT073. Comparing anaerobic genome-scale Fur DNA binding with Fur-dependent transcript expression and protein levels of the uropathogen to that of commensal E. coli K-12 strain MG1655 showed that the Fur regulon of the core genome is conserved but also includes genes within the pathogenicity/genetic islands. Unexpectedly, regulons indicative of amino acid limitation and the general stress response were also indirectly activated in the uropathogen fur mutant, suggesting that induction of the Fur regulon increases amino acid demand. Using RpoS levels as a proxy, addition of amino acids mitigated the stress. In addition, iron chelation increased RpoS to the same levels as in the fur mutant. The increased amino acid demand of the fur mutant or iron chelated cells was exacerbated by aerobic conditions, which could be partly explained by the O2-dependent synthesis of the siderophore aerobactin, encoded by an operon within a pathogenicity island. Taken together, these data suggest that in the iron-poor environment of the urinary tract, amino acid availability could play a role in the proliferation of this uropathogen, particularly if there is sufficient O2 to produce aerobactin. IMPORTANCE Host iron restriction is a common mechanism for limiting the growth of pathogens. We compared the regulatory network controlled by Fur in uropathogenic E. coli (UPEC) to that of nonpathogenic E. coli K-12 to uncover strategies that pathogenic bacteria use to overcome iron limitation. Although iron homeostasis functions were regulated by Fur in the uropathogen as expected, a surprising finding was the activation of the stringent and general stress responses in the uropathogen fur mutant, which was rescued by amino acid addition. This coordinated global response could be important in controlling growth and survival under nutrient-limiting conditions and during transitions from the nutrient-rich environment of the lower gastrointestinal (GI) tract to the more restrictive environment of the urinary tract. The coupling of the response of iron limitation to increased demand for amino acids could be a critical attribute that sets UPEC apart from other E. coli pathotypes.

scholarly journals CBIO-10. REDUCED IRON EXPORT FUNCTIONS IN A CELL INTRINSIC MANNER TO DRIVE GLIOBLASTOMA GROWTH

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii17-ii17
Author(s):  
Katie Troike ◽  
Erin Mulkearns-Hubert ◽  
Daniel Silver ◽  
James Connor ◽  
Justin Lathia
Keyword(s):  
Tumor Cells ◽  
Iron Uptake ◽  
Iron Homeostasis ◽  
Iron Status ◽  
Tumor Grade ◽  
Hfe Gene ◽  
Iron Release ◽  
Female Patients ◽  
Cellular Iron

Abstract Glioblastoma (GBM), the most common primary malignant brain tumor in adults, is characterized by invasive growth and poor prognosis. Iron is a critical regulator of many cellular processes, and GBM tumor cells have been shown to modulate expression of iron-associated proteins to enhance iron uptake from the surrounding microenvironment, driving tumor initiation and growth. While iron uptake has been the central focus of previous investigations, additional mechanisms of iron regulation, such as compensatory iron efflux, have not been explored in the context of GBM. The hemochromatosis (HFE) gene encodes a transmembrane glycoprotein that aids in iron homeostasis by limiting cellular iron release, resulting in a sequestration phenotype. We find that HFE is upregulated in GBM tumors compared to non-tumor brain and that expression of HFE increases with tumor grade. Furthermore, HFE mRNA expression is associated with significantly reduced survival specifically in female patients with GBM. Based on these findings, we hypothesize that GBM tumor cells upregulate HFE expression to augment cellular iron loading and drive proliferation, ultimately leading to reduced survival of female patients. To test this hypothesis, we generated Hfe knockdown and overexpressing mouse glioma cell lines. We observed significant alterations in the expression of several iron handling genes with Hfe knockdown or overexpression, suggesting global disruption of iron homeostasis. Additionally, we show that knockdown of Hfe in these cells increases apoptosis and leads to a significant impairment of tumor growth in vivo. These findings support the hypothesis that Hfe is a critical regulator of cellular iron status and contributes to tumor aggression. Future work will include further exploration of the mechanisms that contribute to these phenotypes as well as interactions with the tumor microenvironment. Elucidating the mechanisms by which iron effulx contributes to GBM may inform the development of next-generation targeted therapies.

scholarly journals Titan Cells Confer Protection from Phagocytosis in Cryptococcus neoformans Infections

2012 ◽  
Vol 11 (6) ◽  
pp. 820-826 ◽  
Author(s):  
Laura H. Okagaki ◽  
Kirsten Nielsen
Keyword(s):  
Cryptococcus Neoformans ◽  
Critical Role ◽  
Cell Formation ◽  
Pulmonary Cryptococcosis ◽  
Content Type ◽  
Cell Production ◽  
Human Fungal Pathogen ◽  
Large Size ◽  
Large Particles ◽  
The Relationship

ABSTRACTThe human fungal pathogenCryptococcus neoformansproduces an enlarged “titan” cell morphology when exposed to the host pulmonary environment. Titan cells exhibit traits that promote survival in the host. Previous studies showed that titan cells are not phagocytosed and that increased titan cell production in the lungs results in reduced phagocytosis of cryptococcal cells by host immune cells. Here, the effect of titan cell production on host-pathogen interactions during early stages of pulmonary cryptococcosis was explored. The relationship between titan cell production and phagocytosis was found to be nonlinear; moderate increases in titan cell production resulted in profound decreases in phagocytosis, with significant differences occurring within the first 24 h of the infection. Not only were titan cells themselves protected from phagocytosis, but titan cell formation also conferred protection from phagocytosis to normal-size cryptococcal cells. Large particles introduced into the lungs were not phagocytosed, suggesting the large size of titan cells protects against phagocytosis. The presence of large particles was unable to protect smaller particles from phagocytosis, revealing that titan cell size alone is not sufficient to provide the observed cross-protection of normal-size cryptococcal cells. These data suggest that titan cells play a critical role in establishment of the pulmonary infection by promoting the survival of the entire population of cryptococcal cells.

scholarly journals Modulation of intestinal sulfur assimilation metabolism regulates iron homeostasis

2018 ◽  
Vol 115 (12) ◽  
pp. 3000-3005 ◽  
Author(s):  
Benjamin H. Hudson ◽  
Andrew T. Hale ◽  
Ryan P. Irving ◽  
Shenglan Li ◽  
John D. York
Keyword(s):  
Iron Deficiency ◽  
Iron Deficiency Anemia ◽  
Iron Reduction ◽  
Genetic Basis ◽  
Iron Homeostasis ◽  
Deficiency Anemia ◽  
Sulfur Assimilation ◽  
Nucleotide Hydrolysis ◽  
Evolutionarily Conserved ◽  
Organismal Development

Sulfur assimilation is an evolutionarily conserved pathway that plays an essential role in cellular and metabolic processes, including sulfation, amino acid biosynthesis, and organismal development. We report that loss of a key enzymatic component of the pathway, bisphosphate 3′-nucleotidase (Bpnt1), in mice, both whole animal and intestine-specific, leads to iron-deficiency anemia. Analysis of mutant enterocytes demonstrates that modulation of their substrate 3′-phosphoadenosine 5′-phosphate (PAP) influences levels of key iron homeostasis factors involved in dietary iron reduction, import and transport, that in part mimic those reported for the loss of hypoxic-induced transcription factor, HIF-2α. Our studies define a genetic basis for iron-deficiency anemia, a molecular approach for rescuing loss of nucleotidase function, and an unanticipated link between nucleotide hydrolysis in the sulfur assimilation pathway and iron homeostasis.

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