The pathogenic yeast Candida albicans (Ca) is a major cause of fungal infections in immune- compromised populations including AIDS patients. It is usually treated with antifungal drugs, most commonly the azole fluconazole which is used extensively in AIDS patients. In these patients, there is a significant probability that azole resistance will develop. Using oral resistant isolates from AIDS patients, major mechanisms of resistance have been identified including alterations in ERG11 (a gene encoding an enzyme in ergosterol biosynthesis and target of the azoles) and increased expression of efflux pumps. Ergosterol is the major sterol in the fungal plasma membrane; and its biosynthesis is the target for azoles and many other antifungals The interaction of fungal cells with azoles is a complex process. Specific aspects of metabolism and the environment influence drug/cell interactions in vitro and have the potential to be important clinically. The Overall Goal of this research is to understand how a fungal cell responds to azoles. This proposal investigates import, subsequent regulation of sterols by the UPC2 transcription factor, and the influence of environmental factors on the cellular response to drugs.
The Specific Aims of this proposal are: 1. To characterize fluconazole import into the fungal cell. Azole import is important to the cell/drug interaction; it may be mediated by passive or active transport; and it has not been studied previously. 2. To analyze the transcriptional regulation of UPC2. Once within the cell, azoles inhibit Erg 11 p, altering sterol levels, and activating the UPC2 transcription factor, which regulates sterol biosynthesis and uptake. Regulation of the UPC2 gene will be analyzed to understand how changes in sterol metabolism, including uptake and biosynthesis are correlated with expression of UPC2. 3. To characterize the effect of specific environmental factors on drug susceptibility and the cell surface. Environmental factors, including pH, oxygen levels, nitrogen sources, and exogenous sterols, will be analyzed for their effect on the drug susceptibility and the cell surface, using microbiological, biochemical and gene expression approaches. 4. To characterize clinical isolates for alterations in sterol metabolism. The known mechanisms of resistance do not apply to many resistant strains.
This aim will assay import, UPC2 regulation, and response to environmental factors to identify possible new resistance mechanisms in these isolates. Understanding the interactions between azoles and fungal cells is a clinically significant issue, with the potential for improving diagnosis, treatment and prevention of fungal infections and resistance.

Agency
National Institute of Health (NIH)
Institute
National Institute of Dental & Craniofacial Research (NIDCR)
Type
Research Project (R01)
Project #
5R01DE017078-03
Application #
7361366
Study Section
AIDS-associated Opportunistic Infections and Cancer Study Section (AOIC)
Program Officer
Rodriguez-Chavez, Isaac R
Project Start
2006-02-01
Project End
2011-01-31
Budget Start
2008-02-01
Budget End
2009-01-31
Support Year
3
Fiscal Year
2008
Total Cost
$400,317
Indirect Cost
Name
Seattle Biomedical Research Institute
Department
Type
DUNS #
070967955
City
Seattle
State
WA
Country
United States
Zip Code
98109
Bhattacharya, Somanon; Esquivel, Brooke D; White, Theodore C (2018) Overexpression or Deletion of Ergosterol Biosynthesis Genes Alters Doubling Time, Response to Stress Agents, and Drug Susceptibility in Saccharomyces cerevisiae. MBio 9:
Zavrel, Martin; White, Theodore C (2015) Medically important fungi respond to azole drugs: an update. Future Microbiol 10:1355-73
Esquivel, Brooke D; Smith, Adam R; Zavrel, Martin et al. (2015) Azole drug import into the pathogenic fungus Aspergillus fumigatus. Antimicrob Agents Chemother 59:3390-8
Zavrel, Martin; Hoot, Sam J; White, Theodore C (2013) Comparison of sterol import under aerobic and anaerobic conditions in three fungal species, Candida albicans, Candida glabrata, and Saccharomyces cerevisiae. Eukaryot Cell 12:725-38
Song, Min-Hee; Lee, Jang-Won; Kim, Min Su et al. (2012) A flucytosine-responsive Mbp1/Swi4-like protein, Mbs1, plays pleiotropic roles in antifungal drug resistance, stress response, and virulence of Cryptococcus neoformans. Eukaryot Cell 11:53-67
Higgins, J; Pinjon, E; Oltean, H N et al. (2012) Triclosan antagonizes fluconazole activity against Candida albicans. J Dent Res 91:65-70
Brown, Gordon D; Denning, David W; Gow, Neil A R et al. (2012) Hidden killers: human fungal infections. Sci Transl Med 4:165rv13
Hoot, Samantha J; Zheng, Xiuzhong; Potenski, Catherine J et al. (2011) The role of Candida albicans homologous recombination factors Rad54 and Rdh54 in DNA damage sensitivity. BMC Microbiol 11:214
Hoot, Samantha J; Smith, Adam R; Brown, Ryan P et al. (2011) An A643V amino acid substitution in Upc2p contributes to azole resistance in well-characterized clinical isolates of Candida albicans. Antimicrob Agents Chemother 55:940-2
Hoot, Samantha J; Brown, Ryan P; Oliver, Brian G et al. (2010) The UPC2 promoter in Candida albicans contains two cis-acting elements that bind directly to Upc2p, resulting in transcriptional autoregulation. Eukaryot Cell 9:1354-62

Showing the most recent 10 out of 16 publications