Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Acidification of vesicular compartments plays an important role in a number of cellular transport processes, including protein secretion, metal cofactor insertion, glycosylation and pH stability. In the present study, we identify and characterize a component of the vesicular proton pump, Vph1p, to determine its role in the virulence of the AIDS-related fungal pathogen Cryptococcus neoformans. Insertional mutagenesis and plasmid rescue were used to identify the VPH1 gene by screening for mutants defective in laccase activity. Disruption of VPH1 resulted in defects in three virulence factors (capsule production, laccase and urease expression), as well as a growth defect at 37 degrees C, but only a small growth reduction at 30 degrees C. These effects were duplicated by the vacuolar (H+)-ATPase inhibitor bafilomycin A1. Furthermore, the vph1 insertional mutant was also avirulent in a mouse meningo-encephalitis model. Complementation of the insertional mutant with wild-type VPH1 resulted in a recovery of virulence factor expression, normal growth at 37 degrees C and restoration of full virulence. These studies establish the importance of the VPH1 gene and vesicular acidification in the virulence of C. neoformans.
Mol Microbiol 2001 Nov
PMID:Multiple virulence factors of Cryptococcus neoformans are dependent on VPH1. 1173 51

Cryptococcosis is a systemic infection in humans caused by the opportunistic fungal pathogen, Cryptococcus neoformans. The infection usually presents as chronic meningoencephalitis, but infects via the respiratory tract. A polysaccharide capsule is a major virulence factor, which allows the yeast to resist host defenses. However, the essential role of the capsule in allowing it to resist host defenses during the initial lung infection has not been clearly shown. A mutant acapsular C. neoformans strain 602 was complemented with the CAP64 gene to obtain an encapsulated strain, TYCC38-602. TYCC38-602 persisted in the lungs of C.B-17 mice after intratracheal inoculation and disseminated to the brain, whereas the mutant acapsular 602 and the plasmid control transformant CIP3-602 strains grew less readily in the lung and were infrequently detected in the brain. T cell-mediated immunity, developed to the encapsulated organism, was required to control growth within the lungs and had a significant impact on numbers of yeasts detected in the brain. The parent acapsular strain, but not the transformant control, also required T cells for optimal inhibition of growth within the lung, but not for maintaining control of the colony-forming units (cfu) in the brain. In summary, the cryptococcal capsule plays an important role in lung virulence and dissemination to the brain, and intact immunity is required to control lung growth of the encapsulated yeast.
Am J Respir Cell Mol Biol 2002 Mar
PMID:Complementation of a capsule deficient Cryptococcus neoformans with CAP64 restores virulence in a murine lung infection. 1186 34

The bioavailability of Se from raw and cured selenomethionine-enriched (Se-enriched) salmon fillets was assessed in Se-deficient male albino rats (Mol: Wist). A low-Se Torula yeast feed was supplemented with 0, 50, 100, 150 or 200 microg Se/kg as sodium selenite or as Se from raw or cured Se-enriched salmon. The diets were fed to weanling rats for 10 and 30 d. Bioavailability of Se was assessed by metabolic balance, Se accumulation in femur, muscle, liver and plasma, and induction of Se-dependent glutathione peroxidase (EC 1.11.1.9; GSHpx) in plasma as response parameters. Except for the metabolic balance results, the slope-ratio method was used when calculating Se bioavailability from raw or cured Se-enriched fish fillets (test food) relative to sodium selenite (standard). The data for fractional apparent absorption and fractional retention showed differences (P<0.05) among all three Se sources in the order raw salmon > cured salmon > selenite. At 10 d, Se from raw and cured Se-enriched fish fillets tended to be more bioavailable than selenite. This was supported by the observations for Se accumulation in femur and muscle and induction of GSHpx activity. At 30 d, all response parameters showed a higher bioavailability of Se from raw and cured Se-enriched fish fillets compared with selenite. Differences (P<0.05) in Se accumulation in muscle at 10 and 30d, and differences (P<0.05) in fractional apparent absorption and fractional retention suggested that curing salmon altered the utilisation of Se. The experimental results showed that enrichment of fish fillets with selenomethionine yields fillets with high Se bioavailability.
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PMID:Bioavailability of selenium from raw or cured selenomethionine-enriched fillets of Atlantic salmon (Salmo salar) assessed in selenium-deficient rats. 1189 11

The capsule of Cryptococcus neoformans is the most obvious virulence factor of this pathogenic yeast. The main capsule constituents are glucuronoxylomannans (GXM). Although several studies have focused on GXM composition and structure, very little is known about their genetics. To elucidate the relationship between the capsule structure and the pathophysiology of the cryptococcosis, genetic screening for mutant strains producing a structurally modified capsule was set up. Using monoclonal antibodies specific for different capsule sugar epitopes, we isolated strains with different mutated capsule structures (Cas mutants). According to their reactivities with various monoclonal antibodies, the mutants were classified into six groups (Cas1 to Cas6). One Cas2 mutant was used to clone the corresponding gene by complementation. This gene (USX1) encodes the previously identified UDP-xylose synthase. We demonstrated that it is necessary for both capsule xylosylation and C. neoformans virulence.
Mol Microbiol 2002 Aug
PMID:Isolation and characterization of capsule structure mutant strains of Cryptococcus neoformans. 1213 28

Resistance to fluconazole is a possible event during prolonged suppressive drug therapy for cryptococ-cal meningitis, the most frequently encountered life-threatening manifestation of cryptococcosis. The knowledge of this resistance at the molecular level is important for management of cryptococcosis. In order to identify genes involved in azole resistance in Cryptococcus neoformans, a cDNA subtraction library technique was chosen as a strategy. First, a fluconazole-resistant mutant BPY22.17 was obtained from a susceptible clinical isolate BPY22 by in vitro exposure to the drug. Then, a subtractive hybridization procedure was used to compare gene expression between the obtained strains. We identified a cDNA overexpressed in the fluconazole-resistant strain BPY22.17 that was used as a probe to isolate the entire gene in a C. neoformans genomic library. Sequence analysis of this gene identified an ATP Binding Cassette (ABC) transporter-encoding gene called C. neoformans AntiFungal Resistance 1 (CnAFR1). Disruption of CnAFR1 gene in the resistant isolate (BPY22.17) resulted in an enhanced susceptibility of the knock-out mutant cnafr1 against fluconazole, whereas reintroduction of the gene in cnafr1 resulted in restoration of the resistance phenotype, thus confirming that CnAFR1 is involved in fluconazole resistance of C. neoformans. Our findings therefore reveal that an active drug efflux mechanism can be involved in the development of azole resistance in this important human pathogen.
Mol Microbiol 2003 Jan
PMID:Identification and characterization of a Cryptococcus neoformans ATP binding cassette (ABC) transporter-encoding gene, CnAFR1, involved in the resistance to fluconazole. 1251 88

Previous studies have shown that a Deltavph1 Cryptococcus neoformans mutant defective in vesicular acidification lacked several important virulence factors including a copper-containing laccase and was avirulent in a mouse model. In the present studies, we characterized laccase transcription and protein production to obtain insights into the mechanism of the vph1 mutation in this pathogen. Although transcription and protein expression were somewhat reduced, laccase protein was found to be successfully translated and correctly targeted to the cell wall in the Deltavph1 mutant as shown by Western blot and immuno-electron microscopy, despite a complete lack of laccase activity. Laccase activity was substantially restored in metabolically active Deltavph1 cells at 30 degrees C by addition of 100 micro M copper sulphate. This restoration by copper was found to occur through both transcriptional and post-translational mechanisms. Laccase transcriptional induction by copper was found to be dependent on enhancer region II within the 5'-untranslated region of CNLAC1. Copper was also found to restore partial activity to Deltavph1 cells at 0 degrees C, suggesting that cell wall laccase was expressed in the mutant as an apo-enzyme. Apo-laccase restoration by copper was found to be facilitated by an acidic environment, consistent with a role for the vacuolar (H+)-ATPase proton pump in copper assembly of laccase in C. neoformans.
Mol Microbiol 2003 Feb
PMID:Copper-mediated reversal of defective laccase in a Deltavph1 avirulent mutant of Cryptococcus neoformans. 1258 55

The pathogenic yeast Cryptococcus neoformans (Cn) var. gattii causes meningoencephalitis in healthy individuals, unlike the better known Cn varieties grubii and neoformans, which are common in immunocompromised individuals. The virulence determinants and mechanisms of host predilection are poorly defined for var. gattii. The present study focused on the characterization of a Cu,Zn superoxide dismutase (SOD1) gene knock-out mutant constructed by developing a DNA transformation system. The sod1 mutant was highly sensitive to the redox cycling agent menadione, and showed fragmentation of the large vacuole in the cytoplasm, but no other defects were seen in growth, capsule synthesis, mating, sporulation, stationary phase survival or auxotrophies for sulphur-containing amino acids. The sod1 mutant was markedly attenuated in virulence in a mouse model, and it was significantly susceptible to in vitro killing by human neutrophils (PMNs). The deletion of SOD1 also resulted in defects in the expression of a number of virulence factors, i.e. laccase, urease and phospholipase. Complementation of the sod1 mutant with SOD1 resulted in recovery of virulence factor expression and menadione resistance, and in restoration of virulence. Overall, these results suggest that the antioxidant function of Cu,Zn SOD is critical for the pathogenesis of the fungus, but is dispensable in its saprobic life. This report constitutes the first instance in which superoxide dismutase has been directly implicated in the virulence of a fungal pathogen.
Mol Microbiol 2003 Mar
PMID:Characterization of Cu,Zn superoxide dismutase (SOD1) gene knock-out mutant of Cryptococcus neoformans var. gattii: role in biology and virulence. 1262 21

Glycosylphosphatidylinositol (GPI) is a conserved post-translational modification to anchor cell surface proteins to plasma membrane in all eukaryotes. In yeast, GPI mediates cross-linking of cell wall mannoproteins to beta1,6-glucan. We reported previously that the GWT1 gene product is a target of the novel anti-fungal compound, 1-[4-butylbenzyl]isoquinoline, that inhibits cell wall localization of GPI-anchored mannoproteins in Saccharomyces cerevisiae (Tsukahara, K., Hata, K., Sagane, K., Watanabe, N., Kuromitsu, J., Kai, J., Tsuchiya, M., Ohba, F., Jigami, Y., Yoshimatsu, K., and Nagasu, T. (2003) Mol. Microbiol. 48, 1029-1042). In the present study, to analyze the function of the Gwt1 protein, we isolated temperature-sensitive gwt1 mutants. The gwt1 cells were normal in transport of invertase and carboxypeptidase Y but were delayed in transport of GPI-anchored protein, Gas1p, and were defective in its maturation from the endoplasmic reticulum to the Golgi. The incorporation of inositol into GPI-anchored proteins was reduced in gwt1 mutant, indicating involvement of GWT1 in GPI biosynthesis. We analyzed the early steps of GPI biosynthesis in vitro by using membranes prepared from gwt1 and Deltagwt1 cells. The synthetic activity of GlcN-(acyl)PI from GlcN-PI was defective in these cells, whereas Deltagwt1 cells harboring GWT1 gene restored the activity, indicating that GWT1 is required for acylation of inositol during the GPI synthetic pathway. We further cloned GWT1 homologues in other yeasts, Cryptococcus neoformans and Schizosaccharomyces pombe, and confirmed that the specificity of acyl-CoA in inositol acylation, as reported in studies of endogenous membranes (Franzot, S. P., and Doering, T. L. (1999) Biochem. J. 340, 25-32), is due to the properties of Gwt1p itself and not to other membrane components.
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PMID:GWT1 gene is required for inositol acylation of glycosylphosphatidylinositol anchors in yeast. 1271 89

Cell wall integrity is crucial for fungal growth, development and stress survival. In the model yeast Saccharomyces cerevisiae, the cell integrity Mpk1/Slt2 MAP kinase and calcineurin pathways monitor cell wall integrity and promote cell wall remodelling under stress conditions. We have identified the Cryptococcus neoformans homologue of the S. cerevisiae Mpk1/Slt2 MAP kinase and have characterized its role in the maintenance of cell integrity in response to elevated growth temperature and in the presence of cell wall synthesis inhibitors. C. neoformans Mpk1 is required for growth at 37 degrees C in vitro, and this growth defect is suppressed by osmotic stabilization. C. neoformans mutants lacking Mpk1 are attenuated for virulence in the mouse model of cryptococcosis. Phosphorylation of Mpk1 is induced in response to perturbations of cell wall biosynthesis by the antifungal drugs nikkomycin Z (a chitin synthase inhibitor), caspofungin (a beta-1,3-glucan synthase inhibitor), or FK506 (a calcineurin inhibitor), and mutants lacking Mpk1 display enhanced sensitivity to nikkomycin Z and caspofungin. Lastly, we show that calcineurin and Mpk1 play complementing roles in regulating cell integrity in C. neoformans. Our studies demonstrate that pharmacological inhibition of the cell integrity pathway would enhance the activity of antifungal drugs that target the cell wall.
Mol Microbiol 2003 Jun
PMID:The Cryptococcus neoformans MAP kinase Mpk1 regulates cell integrity in response to antifungal drugs and loss of calcineurin function. 1278 63

Cryptococcus neoformans is an opportunistic fungal pathogen with a defined sexual cycle in which the alpha allele of the mating type locus is linked to virulence and haploid differentiation. Here we analysed a conserved MAP kinase cascade composed of mating-type specific (Ste11alpha, Ste12alpha) and non-specific (Ste7, Cpk1) elements. Gene disruption experiments demonstrate that this specialized MAP kinase pathway is required for both mating and cell type-specific differentiation but not for virulence. The Ste11alpha, Ste7 and Cpk1 kinases were found to act as a co-ordinate signalling module, whereas the Ste12alpha transcription factor functions with a redundant partner or in a branched or parallel signalling pathway. Our studies illustrate how MAP kinase cascades can be constructed from cell type-specific and non-specific components, yielding pathways that contribute to cell type-specific patterns of signalling and differentiation.
Mol Microbiol 2003 Jul
PMID:A MAP kinase cascade composed of cell type specific and non-specific elements controls mating and differentiation of the fungal pathogen Cryptococcus neoformans. 1282 43


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