scholarly journals The PbMDJ1 Gene Belongs to a Conserved MDJ1/LON Locus in Thermodimorphic Pathogenic Fungi and Encodes a Heat Shock Protein That Localizes to both the Mitochondria and Cell Wall of Paracoccidioides brasiliensis

2006 ◽  
Vol 5 (2) ◽  
pp. 379-390 ◽  
Author(s):  
Wagner L. Batista ◽  
Alisson L. Matsuo ◽  
Luciane Ganiko ◽  
Tânia F. Barros ◽  
Thiago R. Veiga ◽  
...  
Keyword(s):  
Cell Wall ◽  
Heat Shock ◽  
Cell Surface ◽  
Paracoccidioides Brasiliensis ◽  
Pathogenic Fungus ◽  
Fungal Growth ◽  
Pathogenic Fungi ◽  
Yeast Cells ◽  
Shock Proteins

ABSTRACT J-domain (DnaJ) proteins, of the Hsp40 family, are essential cofactors of their cognate Hsp70 chaperones, besides acting as independent chaperones. In the present study, we have demonstrated the presence of Mdj1, a mitochondrial DnaJ member, not only in the mitochondria, where it is apparently sorted, but also in the cell wall of Paracoccidioides brasiliensis, a thermodimorphic pathogenic fungus. The molecule (PbMdj1) was localized to fungal yeast cells using both confocal and electron microscopy and also flow cytometry. The anti-recombinant PbMdj1 antibodies used in the reactions specifically recognized a single 55-kDa mitochondrial and cell wall (alkaline β-mercaptoethanol extract) component, compatible with the predicted size of the protein devoid of its matrix peptide-targeting signal. Labeling was abundant throughout the cell wall and especially in the budding regions; however, anti-PbMdj1 did not affect fungal growth in the concentrations tested in vitro, possibly due to the poor access of the antibodies to their target in growing cells. Labeled mitochondria stood preferentially close to the plasma membrane, and gold particles were detected in the thin space between them, toward the cell surface. We show that Mdj1 and the mitochondrial proteinase Lon homologues are heat shock proteins in P. brasiliensis and that their gene organizations are conserved among thermodimorphic fungi and Aspergillus, where the genes are adjacent and have a common 5′ region. This is the first time a DnaJ member has been observed on the cell surface, where its function is speculative.

scholarly journals Absence of Interleukin-4 Determines Less Severe Pulmonary Paracoccidioidomycosis Associated with Impaired Th2 Response

2004 ◽  
Vol 72 (4) ◽  
pp. 2369-2378 ◽  
Author(s):  
Adriana Pina ◽  
Rita C. Valente-Ferreira ◽  
Eugênia E. W. Molinari-Madlum ◽  
Celidéia A. C. Vaz ◽  
Alexandre C. Keller ◽  
...  
Keyword(s):  
Paracoccidioides Brasiliensis ◽  
Fungal Growth ◽  
Protective Role ◽  
Yeast Cells ◽  
Interleukin 4 ◽  
Delayed Type Hypersensitivity ◽  
Th2 Response ◽  
Leukocyte Influx ◽  
Ifn Γ

ABSTRACT Host resistance to paracoccidiodomycosis, the main deep mycosis in Latin America, is mainly due to cellular immunity and gamma interferon (IFN-γ) production. To assess the role of interleukin-4 (IL-4), a Th2-inducing cytokine, pulmonary paracoccidioidomycosis was studied in IL-4-deficient (IL-4−/−) and wild-type (WT) C57BL/6 mice at the innate and acquired phases of immune response. Forty-eight hours after infection, equivalent numbers of viable Paracoccidioides brasiliensis yeast cells were recovered from the lungs of IL-4−/− and WT mice intratracheally infected with one million fungal cells. Alveolar macrophages from infected IL-4−/− mice controlled in vitro fungal growth more efficiently than macrophages from WT mice and secreted higher levels of nitric oxide. Compared with WT mice, IL-4−/− animals presented increased levels of pulmonary IFN-γ and augmented polymorphonuclear leukocyte influx to the lungs. Decreased pulmonary fungal loads were characterized in deficient mice at week 2 postinfection, concomitant with diminished presence of IL-10. At week 8, lower numbers of yeasts were recovered from lungs and liver of IL-4−/− mice associated with increased production of IFN-γ but impaired synthesis of IL-5 and IL-10. However, a clear shift to a Th1 pattern was not characterized, since IL-4−/− mice did not alter delayed-type hypersensitivity anergy or IL-2 levels. In addition, IL-4 deficiency resulted in significantly reduced levels of pulmonary IL-12, granulocyte-macrophage colony-stimulating factor, IL-3, monocyte chemotactic protein 1, and specific antibody isotypes. In IL-4−/− mice, well-organized granulomas restraining fungal cells replaced the more extensive lesions containing high numbers of fungi and inflammatory leukocytes developed by IL-4-sufficient mice. These results clearly showed that genetically determined deficiency of IL-4 can exert a protective role in pulmonary paracoccidioidomycosis.

Hydrophobic cell wall protein glycosylation by the pathogenic fungus Candida albicans

1994 ◽  
Vol 40 (4) ◽  
pp. 266-272 ◽  
Author(s):  
Kevin C. Hazen ◽  
Pati M. Glee
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Cell Surface ◽  
Molecular Mass ◽  
Pathogenic Fungus ◽  
Cell Surface Hydrophobicity ◽  
Surface Hydrophobicity ◽  
Protein Glycosylation ◽  
Yeast Cells ◽  
Cell Wall Proteins

Cell surface hydrophobicity influences adhesion and virulence of the opportunistic fungal pathogen Candida albicans. Previous studies have shown that cell surface hydrophobicity is due to specific proteins that are exposed on hydrophobic cells but are masked by long fibrils on hydrophilic cells. This observation suggests that hydrophobic cell wall proteins may contain little or no mannosylation. In the present study, the glycosylation levels of three hydrophobic cell wall proteins (molecular mass range between 36 and 40 kDa) derived from yeast cells were examined. One hydrophilic protein (90 kDa) was also tested. Various endoglycosidases (endoglycosidase F – N-glycosidase F, O-glycosidase, β-mannosidase, N-glycosidase F), an exoglycosidase (α-mannosidase), and trifluoromethane sulfonic acid were used to deglycosylate the proteins. All four proteins were reactive to the lectin concanavalin A, demonstrating that they were mannoproteins. However, gel electrophoresis of the control and treated proteins revealed that mannosyl groups of hydrophobic proteins were less than 2 kDa in size, while the mannosyl group of the hydrophilic protein had a molecular mass of approximately 20 kDa. These results suggest that unlike many hydrophilic proteins, hydrophobic proteins may have low levels of glycosylation. Changes in glycosylation may determine exposure of hydrophobic protein regions at the cell surface.Key words: Candida albicans, cell wall, mannoproteins, hydrophobicity, fibrils.

scholarly journals Mechanism of Growth Regulation of Yeast Involving Hydrogen Sulfide From S-Propargyl-Cysteine Catalyzed by Cystathionine-γ-Lyase

2021 ◽  
Vol 12 ◽  
Author(s):  
Zhongkai Gu ◽  
Yufan Sun ◽  
Feizhen Wu
Keyword(s):  
Hydrogen Sulfide ◽  
Growth Regulation ◽  
Biological Activities ◽  
Fungal Growth ◽  
Pathogenic Fungi ◽  
Signaling Molecules ◽  
Yeast Cells ◽  
Anti Fungal

Pathogenic fungi are recognized as a progressive threat to humans, particularly those with the immunocompromised condition. The growth of fungi is controlled by several factors, one of which is signaling molecules, such as hydrogen sulfide (H2S), which was traditionally regarded as a toxic gas without physiological function. However, recent studies have revealed that H2S is produced enzymatically and endogenously in several species, where it serves as a gaseous signaling molecule performing a variety of critical biological functions. However, the influence of this endogenous H2S on the biological activities occurring within the pathogenic fungi, such as transcriptomic and phenotypic alternations, has not been elucidated so far. Therefore, the present study was aimed to decipher this concern by utilizing S-propargyl-cysteine (SPRC) as a novel and stable donor of H2S and Saccharomyces cerevisiae as a fungal model. The results revealed that the yeast could produce H2S by catabolizing SPRC, which facilitated the growth of the yeast cells. This implies that the additional intracellularly generated H2S is generated primarily from the enhanced sulfur-amino-acid-biosynthesis pathways and serves to increase the growth rate of the yeast, and presumably the growth of the other fungi as well. In addition, by deciphering the implicated pathways and analyzing the in vitro enzymatic activities, cystathionine-γ-lyase (CYS3) was identified as the enzyme responsible for catabolizing SPRC into H2S in the yeast, which suggested that cystathionine-γ-lyase might play a significant role in the regulation of H2S-related transcriptomic and phenotypic alterations occurring in yeast. These findings provide important information regarding the mechanism underlying the influence of the gaseous signaling molecules such as H2S on fungal growth. In addition, the findings provide a better insight to the in vivo metabolism of H2S-related drugs, which would be useful for the future development of anti-fungal drugs.

scholarly journals Biosynthesis of β-(1→5)-Galactofuranosyl Chains of Fungal-Type and O-Mannose-Type Galactomannans within the Invasive Pathogen Aspergillus fumigatus

2019 ◽  
Author(s):  
Yuria Chihara ◽  
Yutaka Tanaka ◽  
Minoru Izumi ◽  
Daisuke Hagiwara ◽  
Akira Watanabe ◽  
...  
Keyword(s):  
Cell Wall ◽  
Cell Surface ◽  
Aspergillus Fumigatus ◽  
Pathogenic Fungus ◽  
Cell Surface Hydrophobicity ◽  
Surface Hydrophobicity ◽  
Wall Structure ◽  
Fungal Cell ◽  
Animal Pathogens

ABSTRACTThe pathogenic fungus Aspergillus fumigatus contains galactomannans localized on the surface layer of its cell walls, which are involved in various biological processes. Galactomannans comprise α-(1→2)-/α-(1→6)-mannan and β-(1→5)-/β-(1→6)-galactofuranosyl chains. We previously revealed that GfsA is a β-galactofuranoside β-(1→5)-galactofuranosyltransferase involved in the biosynthesis of β-(1→5)-galactofuranosyl chains. Here, we clarified the entire biosynthesis of β-(1→5)-galactofuranosyl chains in A. fumigatgus. Two paralogs exist within A. fumigatus: GfsB and GfsC. We show that GfsB and GfsC, in addition to GfsA, are β-galactofuranoside β-(1→5)-galactofuranosyltransferases by biochemical and genetic analyses. GfsA, GfsB, and GfsC can synthesize β-(1→5)-galactofuranosyl oligomers up to lengths of 7, 3, and 5 galactofuranoses within an established in vitro highly efficient assay of galactofuranosyltransferase activity. Structural analyses of galactomannans extracted from the strains ΔgfsB, ΔgfsC, ΔgfsAC, and ΔgfsABC revealed that GfsA and GfsC synthesized all β-(1→5)-galactofuranosyl residues of fungal-type and O-mannose-type galactomannans, and GfsB exhibited limited function in A. fumigatus. The loss of β-(1→5)-galactofuranosyl residues decreased the hyphal growth rate and conidia formation ability as well as increased the abnormal hyphal branching structure and cell surface hydrophobicity, but this loss is dispensable for sensitivity to antifungal agents and virulence toward immune-compromised mice.IMPORTANCEβ-(1→5)-galactofuranosyl residues are widely distributed in the subphylum Pezisomycotina of the phylum Ascomycota. Pezizomycotina includes many plant and animal pathogens. Although the structure of β-(1→5)-galactofuranosyl residues of galactomannans in filamentous fungi was discovered long ago, it remains unclear which enzyme is responsible for biosynthesis of this glycan. Fungal cell wall formation processes are complicated, and information concerning glycosyltransferases is essential for their understanding. In this study, we show that GfsA and GfsC are responsible for the biosynthesis of all β-(1→5)-galactofuranosyl residues of fungal-type and O-mannose-type galactomannans. The data presented here indicates that β-(1→5)-galactofuranosyl residues are involved in cell growth, conidiation, polarity, and cell surface hydrophobicity. Our new understanding of β-(1→5)-galactofuranosyl residue biosynthesis provides important novel insights into the formation of the complex cell wall structure and the virulence of the subphylum Pezisomycotina.

In Vitro Holdase Activity of E. coli Small Heat-Shock Proteins IbpA, IbpB and IbpAB: A Biophysical Study with Some Unconventional Techniques

2014 ◽  
Vol 21 (6) ◽  
pp. 564-571 ◽  
Author(s):  
Sourav Roy ◽  
Monobesh Patra ◽  
Suman Nandy ◽  
Milon Banik ◽  
Rakhi Dasgupta ◽  
...  
Keyword(s):  
Heat Shock ◽  
Heat Shock Proteins ◽  
Small Heat Shock Proteins ◽  
E Coli ◽  
Biophysical Study ◽  
Shock Proteins

Translation of Some Maize Small Heat Shock Proteins Is Initiated From Internal In-Frame AUGs

Genetics ◽  
1998 ◽  
Vol 148 (1) ◽  
pp. 471-477
Author(s):  
J Roger H Frappier ◽  
David B Walden ◽  
Burr G Atkinson
Keyword(s):  
Heat Shock ◽  
Heat Shock Proteins ◽  
Single Gene ◽  
Small Heat Shock Proteins ◽  
Shock Proteins

Abstract Etiolated maize radicles (inbred Oh43) subjected to a brief heat shock synthesize a family of small heat shock proteins (≃18 kD) that is composed of at least 12 members. We previously described the cDNA-derived sequence of three maize shsp mRNAs (cMHSP18-1, cMHSP18-3, and cMHSP18-9). In this report, we demonstrate that the mRNA transcribed in vitro from one of these cDNAs (cMHSP 18-9) is responsible for the synthesis of three members of the shsp family, and we suggest that cMHSP18-3 may be responsible for the synthesis of three additional members and cMHSP18-1 for the synthesis of two other members of this family. The fact that these genes do not contain introns, coupled with the observations reported herein, suggest that maize may have established another method of using a single gene to produce a number of different proteins.

scholarly journals Polyhydroxyalkanoate/Antifungal Polyene Formulations with Monomeric Hydroxyalkanoic Acids for Improved Antifungal Efficiency

Antibiotics ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 737
Author(s):  
Marina Pekmezovic ◽  
Melina Kalagasidis Krusic ◽  
Ivana Malagurski ◽  
Jelena Milovanovic ◽  
Karolina Stępień ◽  
...  
Keyword(s):  
Fungal Growth ◽  
Pathogenic Fungi ◽  
Trichophyton Mentagrophytes ◽  
Microsporum Gypseum ◽  
Medium Chain Length ◽  
Gold Drugs ◽  
Growth Inhibitory Activity ◽  
Transdermal Application ◽  
Antifungal Efficiency

Novel biodegradable and biocompatible formulations of “old” but “gold” drugs such as nystatin (Nys) and amphotericin B (AmB) were made using a biopolymer as a matrix. Medium chain length polyhydroxyalkanoates (mcl-PHA) were used to formulate both polyenes (Nys and AmB) in the form of films (~50 µm). Thermal properties and stability of the materials were not significantly altered by the incorporation of polyenes in mcl-PHA, but polyene containing materials were more hydrophobic. These formulations were tested in vitro against a panel of pathogenic fungi and for antibiofilm properties. The films containing 0.1 to 2 weight % polyenes showed good activity and sustained polyene release for up to 4 days. A PHA monomer, namely 3-hydroxydecanoic acid (C10-OH), was added to the films to achieve an enhanced synergistic effect with polyenes against fungal growth. Mcl-PHA based polyene formulations showed excellent growth inhibitory activity against both Candida yeasts (C. albicans ATCC 1023, C. albicans SC5314 (ATCC MYA-2876), C. parapsilosis ATCC 22019) and filamentous fungi (Aspergillus fumigatus ATCC 13073; Trichophyton mentagrophytes ATCC 9533, Microsporum gypseum ATCC 24102). All antifungal PHA film preparations prevented the formation of a C. albicans biofilm, while they were not efficient in eradication of mature biofilms, rendering them suitable for the transdermal application or as coatings of implants.

Glycoconjugates on the surface of spores of the pathogenic fungus Phytophthora cinnamomi studied using fluorescence and electron microscopy and flow cytometry

1990 ◽  
Vol 36 (3) ◽  
pp. 183-192 ◽  
Author(s):  
A. R. Hardham ◽  
E. Suzaki
Keyword(s):  
Flow Cytometry ◽  
Plasma Membrane ◽  
Cell Surface ◽  
Phytophthora Cinnamomi ◽  
Pathogenic Fungus ◽  
Fluorescein Isothiocyanate ◽  
Pathogenic Fungi ◽  
Surface Flow ◽  
Soybean Agglutinin ◽  
Binding Material

Glycoconjugates on the surface of zoospores and cysts of the pathogenic fungus Phytophthora cinnamomi have been studied using fluorescein isothiocyanate labelled lectins for fluorescence microscopy and flow cytometry, and ferritin- and gold-labelled lectins for ultrastructural analysis. Of the five lectins used, only concanavalin A (ConA) binds to the surface of the zoospores, including the flagella and water expulsion vacuole. This suggests that of accessible saccharides, glucosyl or mannosyl residues predominate on the outer surface of the zoospore plasma membrane. Early in encystment, a system of flat disc-like cisternae, which underlie the zoospore plasma membrane, vesiculate. These and other small peripheral vesicles quickly disappear. After the induction of encystment, ConA is no longer localised close to the plasma membrane but binds to material loosely associated with the cell surface. Quantitative measurements by flow cytometry indicate that the ConA-binding material is gradually lost from the cell surface. The cyst wall is weakly labelled, but the site of germ tube emergence stains intensely. During the first 2 min after the induction of encystment, material that binds soybean agglutinin, Helix pommatia agglutinin, and peanut agglutinin appears on the surface of the fungal cells. The distribution of this material, rich in galactosyl or N-acetyl-D-galactosaminosyl residues, is initially patchy, but by 5 min the material evenly coats most of the cell surface. Labelling of zoospores in which intracellular sites are accessible indicates that the soybean agglutinin binding material is stored in vesicles that lie beneath the plasma membrane. Quantitation of soybean agglutinin labelling shows that maximum binding occurs 2–3 min after the induction of encystment. Key words: cell surface, flow cytometry, lectins, pathogenic fungi, Phytophthora cinnamomi.

scholarly journals The Effect of Oxidized Dopamine on the Structure and Molecular Chaperone Function of the Small Heat-Shock Proteins, αB-Crystallin and Hsp27

2021 ◽  
Vol 22 (7) ◽  
pp. 3700
Author(s):  
Junna Hayashi ◽  
Jennifer Ton ◽  
Sparsh Negi ◽  
Daniel E. K. M. Stephens ◽  
Dean L. Pountney ◽  
...  
Keyword(s):  
Heat Shock ◽  
Protein Aggregation ◽  
Molecular Chaperone ◽  
Cross Linking ◽  
Αb Crystallin ◽  
The Cross ◽  
Shock Proteins ◽  
And Function

Oxidation of the neurotransmitter, dopamine (DA), is a pathological hallmark of Parkinson’s disease (PD). Oxidized DA forms adducts with proteins which can alter their functionality. αB-crystallin and Hsp27 are intracellular, small heat-shock molecular chaperone proteins (sHsps) which form the first line of defense to prevent protein aggregation under conditions of cellular stress. In vitro, the effects of oxidized DA on the structure and function of αB-crystallin and Hsp27 were investigated. Oxidized DA promoted the cross-linking of αB-crystallin and Hsp27 to form well-defined dimer, trimer, tetramer, etc., species, as monitored by SDS-PAGE. Lysine residues were involved in the cross-links. The secondary structure of the sHsps was not altered significantly upon cross-linking with oxidized DA but their oligomeric size was increased. When modified with a molar equivalent of DA, sHsp chaperone functionality was largely retained in preventing both amorphous and amyloid fibrillar aggregation, including fibril formation of mutant (A53T) α-synuclein, a protein whose aggregation is associated with autosomal PD. In the main, higher levels of sHsp modification with DA led to a reduction in chaperone effectiveness. In vivo, DA is sequestered into acidic vesicles to prevent its oxidation and, intracellularly, oxidation is minimized by mM levels of the antioxidant, glutathione. In vitro, acidic pH and glutathione prevented the formation of oxidized DA-induced cross-linking of the sHsps. Oxidized DA-modified αB-crystallin and Hsp27 were not cytotoxic. In a cellular context, retention of significant chaperone functionality by mildly oxidized DA-modified sHsps would contribute to proteostasis by preventing protein aggregation (particularly of α-synuclein) that is associated with PD.

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