Candida albicans is an important human fungal pathogen that causes superficial mucosal and lethal systemic infections. Prophylactic antifungal treatments often result in the appearance of acquired drug resistance. C. albicans does not undergo meiosis;genetic diversity arises through somatic mitotic events such as chromosome non-disjunction. We recently found that acquired resistance to fluconazole is often conferred by genome rearrangements involving the centromere of chromosome 5. Centromeres (CENs) are the DNA regions where proteins assemble to form kinetochores, the structures that tether chromosomes to the mitotic spindle. They are critical to proper chromosome segregation and ultimately to genome stability and cell survival. C. albicans has regional CENs like those of higher organisms including humans, and unlike the well characterized point centromeres of Saccharomyces cerevisiae. Importantly, C. albicans CENs are much smaller and simpler than the smallest characterized regional CENs, thus providing a unique opportunity to use C. albicans as a model for the study of regional CENs. Our preliminary results show that deletion of a CEN region can be accompanied by formation of a neocentromere, an ectopic centromere that forms at non- centromeric DNA. Furthermore, we have constructed CEN-plasmids that will be useful for many applications. Our long term goals are: a) to understand the DNA and proteins that specify C. albicans centromere function;b) to design useful molecular tools that exploit our understanding of centromere function and facilitate the study of basic and applied processes in C. albicans. Specifically, we will develop strains that use ADE2-marked CEN-plasmids in a powerful screen for compounds with potential therapeutic value. Our work is critical for the development of two types of potential therapies directed at centromeres: drugs that kill fungi by directly targeting essential fungal centromere components that differ from those in the human centromere;and companion therapies that eliminate aneuploidies and therefore could be administered together with currently available antifungals in order to extend their usefulness. We propose to: 1) determine the requirements for CEN function within native chromosomes;2) determine the requirements for the establishment and maintenance of CEN function on CEN-plasmids;and 3) use CEN-plasmids to screen for candidate antifungal drugs and companion drugs that affect chromosome stability and the acquisition of drug resistance and could extend the usefulness of existing antifungal drugs.

Public Health Relevance

Fungal infections are a serious health problem due to the limited number of antifungal drugs available and the rapid acquisition of resistance to antifungals seen in the clinic. In Candida albicans, the most prevalent fungal pathogen of humans, acquired drug resistance arises from defects in chromosome segregation. We will develop tools to study this process and will identify companion drugs that inhibit these defects, thereby extending the usefulness of available antifungal therapies.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI075096-04
Application #
8073958
Study Section
Pathogenic Eukaryotes Study Section (PTHE)
Program Officer
Duncan, Rory A
Project Start
2008-06-01
Project End
2013-05-31
Budget Start
2011-06-01
Budget End
2012-05-31
Support Year
4
Fiscal Year
2011
Total Cost
$561,148
Indirect Cost
Name
University of Minnesota Twin Cities
Department
Genetics
Type
Schools of Medicine
DUNS #
555917996
City
Minneapolis
State
MN
Country
United States
Zip Code
55455
Berman, Judith; Forche, Anja (2017) Haplotyping a Non-meiotic Diploid Fungal Pathogen Using Induced Aneuploidies and SNP/CGH Microarray Analysis. Methods Mol Biol 1551:131-146
Freire-Benéitez, Verónica; Price, R Jordan; Tarrant, Daniel et al. (2016) Candida albicans repetitive elements display epigenetic diversity and plasticity. Sci Rep 6:22989
Anderson, Matthew Z; Wigen, Lauren J; Burrack, Laura S et al. (2015) Real-Time Evolution of a Subtelomeric Gene Family in Candida albicans. Genetics 200:907-19
McCoy, Kelsey M; Tubman, Emily S; Claas, Allison et al. (2015) Physical limits on kinesin-5-mediated chromosome congression in the smallest mitotic spindles. Mol Biol Cell 26:3999-4014
Gerstein, Aleeza C; Berman, Judith (2015) Shift and adapt: the costs and benefits of karyotype variations. Curr Opin Microbiol 26:130-6
Hirakawa, Matthew P; Martinez, Diego A; Sakthikumar, Sharadha et al. (2015) Genetic and phenotypic intra-species variation in Candida albicans. Genome Res 25:413-25
Tsai, Hung-Ji; Baller, Joshua A; Liachko, Ivan et al. (2014) Origin replication complex binding, nucleosome depletion patterns, and a primary sequence motif can predict origins of replication in a genome with epigenetic centromeres. MBio 5:e01703-14
Anderson, Matthew Z; Gerstein, Aleeza C; Wigen, Lauren et al. (2014) Silencing is noisy: population and cell level noise in telomere-adjacent genes is dependent on telomere position and sir2. PLoS Genet 10:e1004436
Haran, John; Boyle, Hannah; Hokamp, Karsten et al. (2014) Telomeric ORFs (TLOs) in Candida spp. Encode mediator subunits that regulate distinct virulence traits. PLoS Genet 10:e1004658
Gerami-Nejad, Maryam; Zacchi, Lucia F; McClellan, Mark et al. (2013) Shuttle vectors for facile gap repair cloning and integration into a neutral locus in Candida albicans. Microbiology 159:565-579

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