scholarly journals The Small GTPase RacA Mediates Intracellular Reactive Oxygen Species Production, Polarized Growth, and Virulence in the Human Fungal Pathogen Aspergillus fumigatus

2010 ◽  
Vol 10 (2) ◽  
pp. 174-186 ◽  
Author(s):  
Haiyan Li ◽  
Bridget M. Barker ◽  
Nora Grahl ◽  
Srisombat Puttikamonkul ◽  
Jeremey D. Bell ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Aspergillus Fumigatus ◽  
Invasive Pulmonary Aspergillosis ◽  
Reactive Oxygen ◽  
Small Gtpase ◽  
Oxygen Species ◽  
Wild Type ◽  
Content Type ◽  
Diphenylene Iodonium

ABSTRACTAspergillus fumigatusis the predominant mold pathogen in immunocompromised patients. In this study, we present the first characterization of the small GTPase RacA inA. fumigatus. To gain insight into the function ofracAin the growth and pathogenesis ofA. fumigatus, we constructed a strain that lacks a functionalracAgene. The ΔracAstrain showed significant morphological defects, including a reduced growth rate and abnormal conidiogenesis on glucose minimal medium. In the ΔracAstrain, apical dominance in the leading hyphae is lost and, instead, multiple axes of polarity emerge. Intriguingly, superoxide production at the hyphal tips was reduced by 25% in the ΔracAstrain. Treatment of wild-type hyphae with diphenylene iodonium, an inhibitor of NADPH oxidase, resulted in phenotypes similar to that of the ΔracAstrain. These data suggest that ΔracAstrain phenotypes may be due to a reduction or alteration in the production of reactive oxygen species. Most surprisingly, despite these developmental and growth abnormalities, the ΔracAstrain retained at least wild-type virulence in both an insect model and two immunologically distinct murine models of invasive pulmonary aspergillosis. These results demonstrate thatin vitrogrowth phenotypes do not always correlate within vivovirulence and raise intriguing questions about the role of RacA inAspergillusvirulence.

scholarly journals Superoxide Dismutase Influences the Virulence of Cryptococcus neoformans by Affecting Growth within Macrophages

2003 ◽  
Vol 71 (1) ◽  
pp. 173-180 ◽  
Author(s):  
Gary M. Cox ◽  
Thomas S. Harrison ◽  
Henry C. McDade ◽  
Carlos P. Taborda ◽  
Garrett Heinrich ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Superoxide Dismutase ◽  
Cryptococcus Neoformans ◽  
Fungal Infections ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Wild Type ◽  
Pathogenic Yeast ◽  
Antioxidant Defense Systems

ABSTRACT Superoxide dismutase (SOD) is an enzyme that converts superoxide radicals into hydrogen peroxide and molecular oxygen and has been shown to contribute to the virulence of many human-pathogenic bacteria through its ability to neutralize toxic levels of reactive oxygen species generated by the host. SOD has also been speculated to be important in the pathogenesis of fungal infections, but the role of this enzyme has not been rigorously investigated. To examine the contribution of SOD to the pathogenesis of fungal infections, we cloned the Cu,Zn SOD-encoding gene (SOD1) from the human-pathogenic yeast Cryptococcus neoformans and made mutants via targeted disruption. The sod1 mutant strains had marked decreases in SOD activity and were strikingly more susceptible to reactive oxygen species in vitro. A sod1 mutant was significantly less virulent than the wild-type strain and two independent reconstituted strains, as measured by cumulative survival in the mouse inhalational model. In vitro studies established that the sod1 strain had attenuated growth compared to the growth of the wild type and a reconstituted strain inside macrophages producing reduced amounts of nitric oxide. These findings demonstrate that (i) the Cu,Zn SOD contributes to virulence but is not required for pathogenicity in C. neoformans; (ii) the decreased virulence of the sod1 strain may be due to increased susceptibility to oxygen radicals within macrophages; and (iii) other antioxidant defense systems in C. neoformans can compensate for the loss of the Cu,Zn SOD in vivo.

scholarly journals Proteins Needed to Activate a Transcriptional Response to the Reactive Oxygen Species Singlet Oxygen

mBio ◽  
2013 ◽  
Vol 4 (1) ◽  
Author(s):  
Tae-Wook Nam ◽  
Eva C. Ziegelhoffer ◽  
Rachelle A. S. Lemke ◽  
Timothy J. Donohue
Keyword(s):  
Reactive Oxygen Species ◽  
Stress Response ◽  
Singlet Oxygen ◽  
Rhodobacter Sphaeroides ◽  
Transcriptional Response ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Wild Type ◽  
Content Type ◽  
Σ Factor

ABSTRACT Singlet oxygen (1O2) is a reactive oxygen species generated by energy transfer from one or more excited donors to molecular oxygen. Many biomolecules are prone to oxidation by 1O2, and cells have evolved systems to protect themselves from damage caused by this compound. One way that the photosynthetic bacterium Rhodobacter sphaeroides protects itself from 1O2 is by inducing a transcriptional response controlled by ChrR, an anti-σ factor which releases an alternative sigma factor, σE, in the presence of 1O2. Here we report that induction of σE-dependent gene transcription is decreased in the presence of 1O2 when two conserved genes in the σE regulon are deleted, including one encoding a cyclopropane fatty acid synthase homologue (RSP2144) or one encoding a protein of unknown function (RSP1091). Thus, we conclude that RSP2144 and RSP1091 are each necessary to increase σE activity in the presence of 1O2. In addition, we found that unlike in wild-type cells, where ChrR is rapidly degraded when 1O2 is generated, turnover of this anti-σ factor is slowed when cells lacking RSP2144, RSP1091, or both of these proteins are exposed to 1O2. Further, we demonstrate that the organic hydroperoxide tert-butyl hydroperoxide promotes ChrR turnover in both wild-type cells and mutants lacking RSP2144 or RSP1091, suggesting differences in the ways different types of oxidants increase σE activity. IMPORTANCE Oxygen serves many crucial functions on Earth; it is produced during photosynthesis and needed for other pathways. While oxygen is relatively inert, it can be converted to reactive oxygen species (ROS) that destroy biomolecules, cause disease, or kill cells. When energy is transferred to oxygen, the ROS singlet oxygen is generated. To understand how singlet oxygen impacts cells, we study the stress response to this ROS in Rhodobacter sphaeroides, a bacterium that, like plants, generates this compound as a consequence of photosynthesis. This paper identifies proteins that activate a stress response to singlet oxygen and shows that they act in a specific response to this ROS. The identified proteins are found in many free-living, symbiotic, or pathogenic bacteria that can encounter singlet oxygen in nature. Thus, our findings provide new information about a stress response to a ROS of broad biological, agricultural, and biomedical importance.

scholarly journals Induction of Mitochondrial Reactive Oxygen Species Production by Itraconazole, Terbinafine, and Amphotericin B as a Mode of Action against Aspergillus fumigatus

2017 ◽  
Vol 61 (11) ◽  
Author(s):  
Elena Shekhova ◽  
Olaf Kniemeyer ◽  
Axel A. Brakhage
Keyword(s):  
Reactive Oxygen Species ◽  
Lipid Peroxidation ◽  
Aspergillus Fumigatus ◽  
Amphotericin B ◽  
Complex I ◽  
Reactive Oxygen ◽  
Antifungal Compounds ◽  
Ros Production ◽  
Oxygen Species ◽  
Content Type

ABSTRACT Drug resistance in fungal pathogens is of incredible importance to global health, yet the mechanisms of drug action remain only loosely defined. Antifungal compounds have been shown to trigger the intracellular accumulation of reactive oxygen species (ROS) in human-pathogenic yeasts, but the source of those ROS remained unknown. In the present study, we examined the role of endogenous ROS for the antifungal activity of the three different antifungal substances itraconazole, terbinafine, and amphotericin B, which all target the fungal cell membrane. All three antifungals had an impact on fungal redox homeostasis by causing increased intracellular ROS production. Interestingly, the elevated ROS levels induced by antifungals were abolished by inhibition of the mitochondrial respiratory complex I with rotenone. Further, evaluation of lipid peroxidation using the thiobarbituric acid assay revealed that rotenone pretreatment decreased ROS-induced lipid peroxidation during incubation of Aspergillus fumigatus with itraconazole and terbinafine. By applying the mitochondrion-specific lipid peroxidation probe MitoPerOx, we also confirmed that ROS are induced in mitochondria and subsequently cause significant oxidation of mitochondrial membrane in the presence of terbinafine and amphotericin B. To summarize, our study suggests that the induction of ROS production contributes to the ability of antifungal compounds to inhibit fungal growth. Moreover, mitochondrial complex I is the main source of deleterious ROS production in A. fumigatus challenged with antifungal compounds.

scholarly journals Candida albicans Induces Arginine Biosynthetic Genes in Response to Host-Derived Reactive Oxygen Species

2012 ◽  
Vol 12 (1) ◽  
pp. 91-100 ◽  
Author(s):  
Claudia Jiménez-López ◽  
John R. Collette ◽  
Kimberly M. Brothers ◽  
Kelly M. Shepardson ◽  
Robert A. Cramer ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Candida Albicans ◽  
Amino Acid ◽  
Reactive Oxygen ◽  
Single Amino Acid ◽  
Dependent Manner ◽  
Oxygen Species ◽  
Biosynthetic Genes ◽  
Content Type

ABSTRACTThe interaction ofCandida albicanswith phagocytes of the host's innate immune system is highly dynamic, and its outcome directly impacts the progression of infection. While the switch to hyphal growth within the macrophage is the most obvious physiological response, much of the genetic response reflects nutrient starvation: translational repression and induction of alternative carbon metabolism. Changes in amino acid metabolism are not seen, with the striking exception of arginine biosynthesis, which is upregulated in its entirety during coculture with macrophages. Using single-cell reporters, we showed here that arginine biosynthetic genes are induced specifically in phagocytosed cells. This induction is lower in magnitude than during arginine starvationin vitroand is driven not by an arginine deficiency within the phagocyte but instead by exposure to reactive oxygen species (ROS). Curiously, these genes are induced in a narrow window of sublethal ROS concentrations.C. albicanscells phagocytosed by primary macrophages deficient in thegp91phoxsubunit of the phagocyte oxidase do not express theARGpathway, indicating that the induction is dependent on the phagocyte oxidative burst.C. albicans argpathway mutants are retarded in germ tube and hypha formation within macrophages but are not notably more sensitive to ROS. We also find that theARGpathway is regulated not by the general amino acid control response but by transcriptional regulators similar to theSaccharomyces cerevisiaeArgR complex. In summary, phagocytosis induces this single amino acid biosynthetic pathway in an ROS-dependent manner.

scholarly journals Hyperthermia Sensitizes Rhizopus oryzae to Posaconazole and Itraconazole Action through Apoptosis

2013 ◽  
Vol 57 (9) ◽  
pp. 4360-4368 ◽  
Author(s):  
Fazal Shirazi ◽  
Michael A. Pontikos ◽  
Thomas J. Walsh ◽  
Nathaniel Albert ◽  
Russell E. Lewis ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Rhizopus Oryzae ◽  
Reactive Oxygen ◽  
Annexin V ◽  
Intracellular Reactive Oxygen Species ◽  
Oxygen Species ◽  
Content Type ◽  
Dependent Inhibition ◽  
Dose Dependent Inhibition

ABSTRACTThe high mortality rate of mucormycosis with currently available monotherapy has created interest in studying novel strategies for antifungal agents. With the exception of amphotericin B (AMB), the triazoles (posaconazole [PCZ] and itraconazole [ICZ]) are fungistaticin vitroagainstRhizopus oryzae. We hypothesized that growth at a high temperature (42°C) results in fungicidal activity of PCZ and ICZ that is mediated through apoptosis.R. oryzaehad high MIC values for PCZ and ICZ (16 to 64 μg/ml) at 25°C; in contrast, the MICs for PCZ and ICZ were significantly lower at 37°C (8 to 16 μg/ml) and 42°C (0.25 to 1 μg/ml). Furthermore, PCZ and ICZ dose-dependent inhibition of germination was more pronounced at 42°C than at 37°C. In addition, intracellular reactive oxygen species (ROS) increased significantly when fungi were exposed to antifungals at 42°C. Characteristic cellular changes of apoptosis inR. oryzaewere induced by the accumulation of intracellular reactive oxygen species. Cells treated with PCZ or ICZ in combination with hyperthermia (42°C) exhibited characteristic markers of early apoptosis: phosphatidylserine externalization visualized by annexin V staining, membrane depolarization visualized by bis-[1,3-dibutylbarbituric acid] trimethine oxonol (DiBAC) staining, and increased metacaspase activity. Moreover, terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL) assay and DAPI (4′,6-diamidino-2-phenylindole) staining demonstrated DNA fragmentation and condensation, respectively. The addition ofN-acetylcysteine increased fungal survival, prevented apoptosis, reduced ROS accumulation, and decreased metacaspase activation. We concluded that hyperthermia, either alone or in the presence of PCZ or ICZ, induces apoptosis inR. oryzae. Local thermal delivery could be a therapeutically useful adjunct strategy for these refractory infections.

scholarly journals Functional Analysis of theTrichoderma harzianum nox1Gene, Encoding an NADPH Oxidase, Relates Production of Reactive Oxygen Species to Specific Biocontrol Activity against Pythium ultimum

2011 ◽  
Vol 77 (9) ◽  
pp. 3009-3016 ◽  
Author(s):  
M. Montero-Barrientos ◽  
R. Hermosa ◽  
R. E. Cardoza ◽  
S. Gutiérrez ◽  
E. Monte
Keyword(s):  
Reactive Oxygen Species ◽  
Reactive Oxygen ◽  
Pythium Ultimum ◽  
Eukaryotic Cells ◽  
Ros Production ◽  
Oxygen Species ◽  
Wild Type ◽  
Nadph Oxidases ◽  
Content Type ◽  
Biocontrol Activity

ABSTRACTThe synthesis of reactive oxygen species (ROS) is one of the first events following pathogenic interactions in eukaryotic cells, and NADPH oxidases are involved in the formation of such ROS. Thenox1gene ofTrichoderma harzianumwas cloned, and its role in antagonism against phytopathogens was analyzed innox1-overexpressed transformants. The increased levels ofnox1expression in these transformants were accompanied by an increase in ROS production during their direct confrontation withPythium ultimum. The transformants displayed an increased hydrolytic pattern, as determined by comparing protease, cellulase, and chitinase activities with those for the wild type. In confrontation assays againstP. ultimumthenox1-overexpressed transformants were more effective than the wild type, but not in assays againstBotrytis cinereaorRhizoctonia solani. A transcriptomic analysis using aTrichodermahigh-density oligonucleotide (HDO) microarray also showed that, compared to gene expression for the interaction of wild-typeT. harzianumandP. ultimum, genes related to protease, cellulase, and chitinase activities were differentially upregulated in the interaction of anox1-overexpressed transformant with this pathogen. Our results show thatnox1is involved inT. harzianumROS production and antagonism againstP. ultimum.

scholarly journals Peroxiredoxin Asp f3 Is Essential for Aspergillus fumigatus To Overcome Iron Limitation during Infection

mBio ◽  
2021 ◽  
Author(s):  
Victor Brantl ◽  
Jana M. Boysen ◽  
Annie Yap ◽  
Evgeny Golubtsov ◽  
Dominik Ruf ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Aspergillus Fumigatus ◽  
Virulence Factor ◽  
Vaccine Candidate ◽  
Reactive Oxygen ◽  
Iron Limitation ◽  
Oxygen Species ◽  
Content Type ◽  
Ros Scavenger ◽  
Fungal Allergen

Asp f3 is one of the most abundant proteins in the pathogenic mold Aspergillus fumigatus . It has an enigmatic multifaceted role as a fungal allergen, virulence factor, reactive oxygen species (ROS) scavenger, and vaccine candidate.

scholarly journals Cdc42GAP, reactive oxygen species, and the vimentin network

2009 ◽  
Vol 297 (2) ◽  
pp. C299-C309 ◽  
Author(s):  
Qing-Fen Li ◽  
Amy M. Spinelli ◽  
Dale D. Tang
Keyword(s):  
Reactive Oxygen Species ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Reactive Oxygen ◽  
Muscle Cells ◽  
Oxygen Species ◽  
Wild Type ◽  
Vitro Assay ◽  
Contractile Activation

Cdc42GAP (GTPase-activating protein) has been implicated in the regulation of cell motility, adhesion, proliferation, and apoptosis. In this study, Cdc42GAP was cloned from smooth muscle tissues. Cdc42GAP, but not inactive R282A Cdc42GAP (alanine substitution at arginine-282), enhanced the GTP hydrolysis of Cdc42 in an in vitro assay. Furthermore, we developed an assay to evaluate the activity of Cdc42GAP in vivo. Stimulation of smooth muscle cells with 5-hydroxytryptamine (5-HT) resulted in the decrease in Cdc42GAP activity. The agonist-induced GAP suppression was reversed by reactive oxygen species inhibitors. Treatment with hydrogen peroxide also inhibited GAP activity in smooth muscle cells. Because the vimentin cytoskeleton undergoes dynamic changes in response to contractile activation, we evaluated the role of Cdc42GAP in regulating vimentin filaments. Smooth muscle cells were infected with retroviruses encoding wild-type Cdc42GAP or its R282A mutant. Expression of wild-type Cdc42GAP, but not mutant R282A GAP, inhibited the increase in the activation of Cdc42 upon agonist stimulation. Phosphorylation of p21-activated kinase (PAK) at Thr-423 (an indication of PAK activation), vimentin phosphorylation (Ser-56), partial disassembly and spatial remodeling, and contraction were also attenuated in smooth muscle cells expressing Cdc42GAP. Our results suggest that the activity of Cdc42GAP is regulated upon contractile activation, which is mediated by intracellular ROS. Cdc42GAP regulates the vimentin network through the Cdc42-PAK pathway in smooth muscle cells during 5-HT stimulation.

scholarly journals Antifungal Activity of Plasmacytoid Dendritic Cells against Cryptococcus neoformansIn VitroRequires Expression of Dectin-3 (CLEC4D) and Reactive Oxygen Species

2016 ◽  
Vol 84 (9) ◽  
pp. 2493-2504 ◽  
Author(s):  
Camaron R. Hole ◽  
Chrissy M. Leopold Wager ◽  
Andrew S. Mendiola ◽  
Karen L. Wozniak ◽  
Althea Campuzano ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Dendritic Cells ◽  
Pulmonary Infection ◽  
Reactive Oxygen ◽  
Optimal Growth ◽  
Plasmacytoid Dendritic Cells ◽  
Oxygen Species ◽  
Content Type ◽  
Lectin Receptor

Conventional dendritic cells (cDCs) are critical for protection against pulmonary infection with the opportunistic fungal pathogenCryptococcus neoformans; however, the role of plasmacytoid dendritic cells (pDCs) is unknown. We show for the first time that murine pDCs have direct activity againstC. neoformansvia reactive oxygen species (ROS), a mechanism different from that employed to controlAspergillus fumigatusinfections. The anticryptococcal activity of murine pDCs is independent of opsonization but appears to require the C-type lectin receptor Dectin-3, a receptor not previously evaluated during cryptococcal infections. Human pDCs can also inhibit cryptococcal growth by a mechanism similar to that of murine pDCs. Experimental pulmonary infection of mice with aC. neoformansstrain that induces protective immunity demonstrated that recruitment of pDCs to the lungs is CXCR3 dependent. Taken together, our results show that pDCs inhibitC. neoformansgrowthin vitrovia the production of ROS and that Dectin-3 is required for optimal growth-inhibitory activity.

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