http://hdl.handle.net/10033/190334 Mon, 20 May 2013 18:54:50 GMT 2013-05-20T18:54:50Z http://hdl.handle.net/10033/283773 Title: Involvement of the mitogen activated protein kinase Hog1p in the response of Candida albicans to iron availability. Authors: Kaba, Hani Ej; Nimtz, Manfred; Müller, Peter P; Bilitewski, Ursula Abstract: ABSTRACT: BACKGROUND: Iron is an essential nutrient for almost all organisms, and generating iron limiting conditions for pathogens is one of the host defense strategies against microbial infections. Excess of iron can be toxic; therefore, iron uptake is tightly controlled. The high affinity iron uptake system of the opportunistic pathogenic yeast Candida albicans has been shown to be essential for virulence. Several transcription factors and regulators of iron uptake genes were identified, but the knowledge of signaling pathways is still limited. Gene expression profiling of the Deltahog1 deletion mutant indicated an involvement of the mitogen activated protein (MAP) kinase Hog1p. However, the function of Hog1p in the response of C. albicans to iron availability was not studied in detail. Thus, we analyzed phenotypic and molecular responses of C. albicans to different iron concentrations particularly with respect to the activity of the Hog1p MAP kinase module. RESULTS: We observed flocculation of yeast cells, when the iron ion concentration was equal to or higher than 5 muM. This phenotype was dependent on the MAP kinase Hog1p and the corresponding MAP kinase kinase Pbs2p. Moreover, high extracellular iron ion concentrations led to hyper-phosphorylation of Hog1p. We determined lower amounts of multicopper ferroxidase (MCFO) proteins and lower ferric reductase activity, when the iron ion concentration in the medium was increased. This effect was also observed for the Deltahog1 mutant. However, the amounts of MCFO proteins and the cell surface ferric reductase activity were increased in the Deltahog1 in comparison to wild type cells. This effect was independent of iron availability in growth media. CONCLUSIONS: In C. albicans, the MAP kinase Hog1p is part of the network regulating the response of the organism to iron availability. Hog1p was transiently phosphorylated under high iron concentrations and was essential for a flocculent phenotype. Furthermore, deletion of HOG1 led to increased levels of components of the reductive iron uptake system in comparison to the wild-type, independent of iron concentrations in the media. However, the additional induction of this system by low iron concentrations was independent of HOG1. Thu, 24 Jan 2013 00:00:00 GMT http://hdl.handle.net/10033/283773 2013-01-24T00:00:00Z http://hdl.handle.net/10033/269716 Title: The fungicide fludioxonil antagonizes fluconazole activity in the human fungal pathogen Candida albicans. Authors: Buschart, Anna; Burakowska, Anna; Bilitewski, Ursula Abstract: The fungicide fludioxonil is widely used in agriculture. Residua of this fungicide are occasionally detected in fruits and can therefore be ingested by humans. The human fungal pathogen Candida albicans expresses the target of fludioxonil, Nik1p, a type III histidine kinase involved in stress response. Inhibition of yeast and hyphae growth was hardly observable after treatment of C. albicans SC5314 with fludioxonil. As a side effect, however, we observed a concentration-dependent induction of the expression of the genes CDR1 and CDR2, encoding ATP-binding cassette (ABC) transporters. This was independent of the presence of the target of fludioxonil as induction was also observed in a Δnik1 deletion mutant. Deletion of the CDR1 gene aggravated the inhibition of germ tube formation by fludioxonil, indicating that, in the wild-type, the fungicide was discharged from the cell by Cdr1p. Cdr1p is also known as a resistance factor of C. albicans against the commonly used antimycotic fluconazole. Thus, the effect of concurrent exposure to fludioxonil and known cargoes of ABC transporters on their extrusion and the growth of C. albicans was examined. Pre-incubation with fludioxonil decreased the export rate of rhodamine 6G. The resistance to fluconazole was increased by fludioxonil, independently of Nik1p. Therefore, exposure of C. albicans to fludioxonil may lead to increased resistance to fluconazole treatment. Sat, 01 Dec 2012 00:00:00 GMT http://hdl.handle.net/10033/269716 2012-12-01T00:00:00Z http://hdl.handle.net/10033/216297 Title: A viability assay for Candida albicans based on the electron transfer mediator 2,6-dichlorophenolindophenol. Authors: Hassan, Rabeay Y A; Bilitewski, Ursula Abstract: Candida albicans is an opportunistic fungal pathogen with comparably high respiratory activity. Thus, we established a viability test based on 2,6-dichlorophenolindophenol (DCIP), a membrane-permeable electron transfer agent. NADH dehydrogenases catalyze the reduction of DCIP by NADH, and the enzymatic activity can be determined either electrochemically via oxidation reactions of DCIP or photometrically. Among the specific respiratory chain inhibitors, only the complex I inhibitor rotenone decreased the DCIP signal from C. albicans, leaving residual activity of approximately 30%. Thus, the DCIP-reducing activity of C. albicans was largely dependent on complex I activity. C. albicans is closely related to the complex I-negative yeast Saccharomyces cerevisiae, which had previously been used in DCIP viability assays. Via comparative studies, in which we included the pathogenic complex I-negative yeast Candida glabrata, we could define assay conditions that allow a distinction of complex I-negative and -positive organisms. Basal levels of DCIP turnover by S.cerevisiae and C. glabrata were only 30% of those obtained from C. albicans but could be increased to the C. albicans level by adding glucose. No significant increases were observed with galactose. DCIP reduction rates from C. albicans were not further increased by any carbon source. Thu, 01 Dec 2011 00:00:00 GMT http://hdl.handle.net/10033/216297 2011-12-01T00:00:00Z http://hdl.handle.net/10033/190339 Title: Improving the prediction of Pseudomonas putida mt-2 growth kinetics with the use of a gene expression regulation model of the TOL plasmid Authors: Koutinas, Michalis; Kiparissides, Alexandros; Lam, Ming-Chi; Silva-Rocha, Rafael; Godinho, Miguel; de Lorenzo, Victor; Martins dos Santos, Vitor A.P.; Pistikopoulos, Efstratios N.; Mantalaris, Athanasios Tue, 22 Nov 2011 15:39:44 GMT http://hdl.handle.net/10033/190339 2011-11-22T15:39:44Z