Paul J. Szaniszlo
| Title: | Professor |  Video - PC Mac |
| Education: | Ph.D.: 1967, University of North Carolina; M.A.: 1964, University of North Carolina; B.A.: 1961, Ohio Wesleyan |
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| Postdoc.: | Harvard University | |
| Research: | Fungal cellular development | |
| Office: | NMS 2.314 | |
| Phone: | (512) 471-3384 | |
| E-mail: | pjszaniszlo@mail.utexas.edu | |
| Postal Address: |
The University of Texas at Austin
Molecular Genetics & Microbiology
1 University Station A5000
Austin TX 78712-0162 |
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| Courses taught: | BIO 341 "Biology of Fungi" BIO 329 "Medical Mycology" |
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| Laboratory home page: http://www.sbs.utexas.edu/mycology/ | ||
My laboratory uses the black yeast Wangiella dermatitidis as a model to discover cell wall-related virulence factors among the more than 100 black fungi known to cause human disease. The importance of our studies relates to the knowledge that fungi blackened by the polymerization of dihydroxynaphthalene (DHN) into melanin (DHN-melanin) in their cell walls are among those most resistant to current strategies of antifungal chemotherapeutic intervention. Therefore, the discovery of critical, but vulnerable, steps leading to the synthesis of DHN-melanin and other cell wall components in our model should identify excellent potential targets for the design of antifungal agents. These compounds should be useful not only for the treatment of infections caused by black fungi, but also other less resistant but more common species.
Over the years, members of my research groups have carried out a variety of studies of Wangiella using techniques of cell biology, genetics, physiology and biochemistry aimed at establishing this fungus as a model. In contrast, during the last decade, my group has mostly been engaged in cloning and studying genes and their encoded products involved in cell wall biosynthesis using a variety of newly developed transformation, gene disruption, and gene expression strategies. The focus of these studies has been toward a molecular genetic analysis of the relevance of chitin and melanin biosynthesis to virulence, and of genes that affect the extent and location of their cell wall deposition. Although progress has been made in all these areas, most has involved the four chitin synthase (WdCHS) structural genes we have identified. Our results to date suggest that: i) no single chitin synthase (WdChs) is required for virulence; ii) whereas WdChs1p and WdChs2p are apparently redundant for function at 25oC and viability at 37oC, of the two, WdChs2p is more important to virulence because strains with only WdChs2p are fully virulent; iii) WdChs3p still contributes to virulence when WdChs2p is defective but WdChs1p is intact, because among mutants that grow at 37oC, only those with defects in both WdChs2p and WdChs3p are less virulent. Currently we are attempting to discover the regulatory mechanisms that control the functions of these two zymogens and allow them to contribute to virulence in association with melanin and in the different growth forms of Wangiella growing at temperatures of infection.
Selected Publications
2004 |
Liu, H., S. Kauffman, J. M. Becker and P. J. Szaniszlo. Wangiella (Exophiala) dermatitidis, WdChs5p, a class V chitin synthase is essential for sustained cell growth at temperature of infection. Eukaryot. Cell. 3 (2004) 40-51.
Cheng, Q., K. A. Kinney, C. P. Whitman, and P. J. Szaniszlo. Characterization of two polyketide synthase genes in Exophiala lecanii-corni, a melanized fungus with bioremediation potential. Bioorgan. Chem. 32 (2004) 92-108.
2002 |
Szaniszlo, P. J. Molecular genetic studies of the model dematiaceous pathogen Wangiella dermatitidis. Int. J. Med. Microbiol. 292 (2002) 381-390.
Wang, Q., H. Liu and P. J. Szaniszlo. Compensatory expression of five chitin synthase genes, a response to stress stimuli in Wangiella (Exophiala) dermatitidis, a melanized fungal pathogen of humans. Microbiology 148 (2002) 2811-2817.
Zheng, W. and P. J. Szaniszlo. Characterization of WdChs3p, a Class III chitin synthase of Wangiella (Exophiala) dermatitidis, expressed in Saccharomyces cerevisiae. Med. Mycol. 40 (2002) 283-289.
2001 |
Liu, H., Z. Wang, L. Zheng, M. Hauser, S. Kauffman, J. M. Becker and P. J. Szaniszlo. Relevance of chitin and chitin synthases to virulence in Wangiella (Exophiala) dermatitidis, a model melanized pathogen of humans. p. 463-472. In R. A. A. Muzzarelli (ed.) Chitin Enzymology 2001. Atec Edizioni, Italy.
Wang, Z., L. Zheng, H. Liu, Q. Wang, M. Hauser, S. Kaufman, J. M. Becker, and P. J. Szaniszlo. WdChs2, a class I chitin synthase, together with WdChs3p (class III), contributes to virulence in Wangiella (Exophiala) dermatitidis. Infect. Immun. 69 (2001) 7517-7526.
Woertz, J. R., K. A. Kinney, N. D. P. McIntosh and P. J. Szaniszlo. Removal of toluene in vapor-phase bioreactor containing a strain of the dimorphic black yeast Exophiala lecanii-corni. Biotechnol. Bioeng. 75 (2001) 550-558.
Woertz, J. R., K. A. Kinney, N. D. P. McIntosh, and P. J. Szaniszlo. A fungal vapor-phase bioreactor for removal of nitric oxide from waste gas streams. J. Air Waste Manag. Assoc. 51 (2001) 895-902.
Feng, B., X. Wang, M. Hauser, S. Kauffmann, S. Jentsch, G. Haase, J. M. Becker, and P. J. Szaniszlo. Molecular cloning and characterization of WdPKS1, a gene involved in dihydroxynaphthalene (DHN) melanin biosynthesis and virulence in Wangiella (Exophiala) dermiatitidis. Infect. Immun. 69 (2001) 1781-1794.
2000 |
Wang, Z. and P. J. Szaniszlo.
The Journal of Bacteriology 182 (2000) 874-881.
ABSTRACT: Class III chitin synthases are important for hyphal growth in some filamentous fungi but are not found in yeasts. Using a specific PCR product that encodes a portion of the class III chitin synthase of W. dermatitidis as a probe, we isolated the chitin synthase gene, WdCHS3, from this polymorphic melanized pathogen of humans. Northern blotting showed that WdCHS3 was highly expressed under stress conditions, such as the shift of cells to temperatures commensurate with infection, or to conditions that induce cellular morphogenesis in this fungus. Analysis of the 5' upstream sequence of WdCHS3 provided evidence for a negative regulatory element at between 780 and 1600 bp. Western blotting indicated that the production of the WdChs3p was temperature dependent and temporally regulated. Disruption of WdCHS3 in a wild-type strain and in two temperature-sensitive morphological mutants resulted in significantly reduced chitin synthase activities but did not obviously affect their morphologies, growth rates, chitin contents, or virulence. This paradox suggested that the contributions of the high levels of WdCHS3 gene expression and WdChs3p production in strains subjected to stress reside in unknown or unexamined parts of the life cycle of this ecologically poorly known member of the Fungi Imperfecti. Nonetheless, this report presents the first evidence that transcription of a chitin synthase gene is regulated by a negative regulatory element in its 5' upstream sequence.Expression of a constitutively active Cdc42 homologue promotes development of sclerotic bodies, but represses hyphal growth in the zoopathogenic fungus Wangiella (Exophiala) dermatitidis.
Ye, X. and P. J. Szaniszlo.
The Journal of Bacteriology 182 (2000) 4941-4950.
ABSTRACT: In contrast to the CDC42 homologues of Saccharomyces cerevisiae and Schizosaccharomyces pombe, the WdCDC42 gene in the human pathogenic fungus Wangiella (Exophiala) dermatitidis was found to be nonessential for cell viability. Expression of the constitutively active allele wdcdc42G14V at 37°C induced nonpolarized growth that led to cell enlargement and multiple nucleation. The swollen cells subsequently converted into planate divided bicellular forms or multiply septated sclerotic bodies in post-log phase, when the G14V-altered protein was diminished. The wdcdc42G14V mutation also strongly repressed filamentous growth both in the wild-type strain and in the temperature-sensitive hyphal-form mutant Hf1. In contrast, overexpression of the dominant negative alleles wdcdc42T19N and wdcdc42D120A had no obvious effect on fungal-cell polarization. These results suggested that WdCdc42p plays a unique regulatory role in cellular morphogenesis in W. dermatitidis. Activation of this protein in response to extracellular or intracellular signals seems to commit its yeast-like cells to a phenotype transition that produces sclerotic bodies while repressing hyphal development.
WdPKS1 encodes a polyketide synthase involved in dihydroxynaphthalene- (DHN-) melanin biosynthesis and virulence in Wangiella (Exophiala) dermatitidis.
Feng, B. and P. J. Szaniszlo.
Infection and Immunity (in press).
1999 |
Zheng, L. and P. J. Szaniszlo.
Medical Mycology 37 (1999) 85-96.
ABSTRACT: A genomic clone encoding the Wangiella dermatitidis orotidine monophosphate pyrophosphorylase gene (WdURA5) was isolated by screening a subgenomic plasmid DNA library of this phaeohyphomycotic agent using a PCR amplification product of the gene as a probe. When plasmid DNA containing the cloned WdURA5 gene was introduced by electroporation into a wdura5 auxotrophic recipient strain derived previously by selection with 5-fluoroorotic acid (5-FOA), an apparent gene repair event occurred at high frequency without any evidence of integration of the plasmid DNA. Therefore, the hygromycin B resistance gene (the hph gene) was used as a dominant selective marker for the disruption of WdURA5 to generate a new, more stable, wdura5 auxotrophic strain. Transformation of this strain was then achieved with high efficiency and high frequency by site-specific integration using WdURA5 as a selective marker. To initiate attempts to use this marker repeatedly for multiple chitin synthase (WdCHS) gene disruptions in single strains of W. dermatitidis, a hisG_WdURA5_hisG cassette was constructed and used to disrupt WdCHS2. The WdURA5 gene in the disruptant was then successfully recycled under selection for resistance to 5-FOA.A color-selectable and site-specific integrative transformation system for gene expression studies in the dematiaceous fungus Wangiella (Exophiala) dermatitidis.
Ye, X., B. Feng, and P. J. Szaniszlo.
Current Genetics 36 (1999) 241-247.
ABSTRACT: To explore potential virulence factors in the dematiaceous (melanized) fungus Wangiella dermatitidis, we established a gene expression system with properties of homologous transformation and color identification. Using a polyketide synthase gene (WdPKS1) fragment for targeting, we found that 52% of transformants became albinos easily distinguishable from nonspecific transformants. Southern analysis confirmed that the integrations were at the WdPKS1 locus, which however did not affect transformant growth. With a heterologous promoter, P-glaA, enhanced expression of lacZ was found at 37 °C. Our results indicated that this system allows the efficient production of isogenic strains for gene function analysis in W. dermatitidis.WdCHS4, a homolog of chitin synthase 3 in Saccharomyces cerevisiae, cannot alone support the growth of Wangiella (Exophiala) dermatitidis at temperature of infection.
Wang, Z., L. Zheng and P. J. Szaniszlo.
Infection and Immunity 67 (1999) 6619-6630.
ABSTRACT: By using improved transformation methods for Wangiella dermatitidis, and a cloned fragment of its chitin synthase 4 structural gene (WdCHS4) as a marking sequence, the full-length gene was rescued from the genome of this human pathogenic fungus. The encoded chitin synthase product (WdChs4p) showed high homology with Chs3p of Saccharomyces cerevisiae and other class IV chitin synthases, and Northern blotting showed that WdCHS4 was expressed at constitutive levels under all conditions tested. Reduced chitin content, abnormal yeast clumpiness and budding kinetics, and increased melanin secretion resulted from the disruption of WdCHS4 suggesting that WdChs4p influences cell wall structure, cellular reproduction, and melanin deposition, respectively. However, no significant loss of virulence was detected when the wdchs4 strain was tested in an acute mouse model. Using a wdchs1 wdchs2 wdchs3 triple mutant of W. dermatitidis, which grew poorly but adequately at 25°C, we assayed WdChs4p activity in the absence of activities contributed by its three other WdChs proteins. Maximal activity required trypsin activation, suggesting a zymogenic nature. The activity also had a pH optimum of 7.5, was most stimulated by Mg2+, and was more inhibited by polyoxin D than by nikkomycin Z. Although the WdChs4p activity had a broad temperature optimum between 30 to 45°C in vitro, this activity alone did not support the growth of the wdchs1 wdchs2 wdchs3 triple mutant at 37°C, a temperature commensurate with infection.
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