Gene annotation and drug target discovery in Candida albicans with a tagged transposon mutant collection
Julia Oh1, Eula Fung, Ulrich Schlecht
1Department of Genetics, Stanford University, Palo Alto, California, United States of America.
Abstract:
Candida albicans is the most common human fungal pathogen, causing infections that can be lethal in immunocompromised patients. Although Saccharomyces cerevisiae has been used as a model for C. albicans, it lacks C. albicans' diverse morphogenic forms and is primarily non-pathogenic. Comprehensive genetic analyses that have been instrumental for determining gene function in S. cerevisiae are hampered in C. albicans, due in part to limited resources to systematically assay phenotypes of loss-of-function alleles. Here, we constructed and screened a library of 3633 tagged heterozygous transposon disruption mutants, using them in a competitive growth assay to examine nutrient- and drug-dependent haploinsufficiency. We identified 269 genes that were haploinsufficient in four growth conditions, the majority of which were condition-specific. These screens identified two new genes necessary for filamentous growth as well as ten genes that function in essential processes. We also screened 57 chemically diverse compounds that more potently inhibited growth of C. albicans versus S. cerevisiae. For four of these compounds, we examined the genetic basis of this differential inhibition. Notably, Sec7p was identified as the target of brefeldin A in C. albicans screens, while S. cerevisiae screens with this compound failed to identify this target. We also uncovered a new C. albicans-specific target, Tfp1p, for the synthetic compound 0136-0228. These results highlight the value of haploinsufficiency screens directly in this pathogen for gene annotation and drug target identification.
Insights
This study developed a new method to understand gene function in Candida albicans, a dangerous fungal pathogen. Haploinsufficiency screens identified new drug targets and genes critical for fungal growth and infection.
Area of Science:
- Medical Mycology
- Molecular Biology
- Genetics
Background:
- Candida albicans is a major human fungal pathogen, often lethal in immunocompromised individuals.
- Saccharomyces cerevisiae, a common model organism, has limitations for studying C. albicans due to differences in morphology and pathogenicity.
- Limited genetic tools in C. albicans hinder systematic analysis of gene function.
Purpose of the Study:
- To develop and apply haploinsufficiency screening in C. albicans for gene function analysis.
- To identify genes involved in nutrient and drug responses.
- To discover novel drug targets for C. albicans infections.
Main Methods:
- Construction and screening of a library of 3633 tagged heterozygous transposon disruption mutants.
- Competitive growth assays to assess nutrient- and drug-dependent haploinsufficiency.
- Screening of 57 diverse compounds for differential inhibition of C. albicans versus S. cerevisiae.
Main Results:
- Identified 269 genes exhibiting condition-specific haploinsufficiency.
- Discovered two novel genes essential for filamentous growth and ten genes in vital processes.
- Identified Sec7p as the target of brefeldin A and Tfp1p as a new target for compound 0136-0228 in C. albicans.
Conclusions:
- Haploinsufficiency screening in C. albicans is a valuable approach for gene annotation.
- This method effectively identifies condition-specific gene functions and potential drug targets.
- The study provides new insights into C. albicans pathogenesis and antifungal drug discovery.
