High-Resolution Genome-Wide Occupancy in Candida spp. Using ChEC-seq.

Area of Science:

• Molecular Biology
• Mycology
• Genetics

Background:

• Pathogenic fungi, including Candida albicans and Candida auris, pose significant public health threats due to their persistence and virulence.
• Understanding transcriptional regulation is crucial for identifying therapeutic targets against fungal infections.
• Existing chromatin immunoprecipitation (ChIP)-based methods like ChIP-seq have limitations, including potential artifacts.

Purpose of the Study:

• To introduce and validate a novel PCR-based chromatin endogenous cleavage coupled to high-throughput sequencing (ChEC-seq) method for pathogenic Candida species.
• To determine the genome-wide location of transcriptional regulators without protein-DNA cross-linking or sonication.
• To enable high-resolution mapping of regulatory elements governing fungal fitness and drug resistance.

Main Methods:

• Development of new MNase-tagging plasmids for Candida species.
• Application of chromatin endogenous cleavage (ChEC) procedure coupled with high-throughput sequencing (ChEC-seq).
• Proof-of-concept mapping of the SWI/SNF complex and Nsi1 transcription factor in Candida albicans.

Main Results:

• Successful implementation of ChEC-seq in Candida species, providing a high-resolution binding map of key regulators.
• Demonstrated avoidance of artifacts associated with traditional ChIP methods (epitope masking, hyper-ChIPable euchromatin).
• Generated detailed genomic occupancy data for the SWI/SNF complex and Nsi1 in Candida albicans.

Conclusions:

• The developed ChEC-seq procedure offers a robust and artifact-free method for genome-wide regulator mapping in pathogenic yeasts.
• This technique enhances understanding of transcriptional regulatory circuits controlling fungal fitness, virulence, and antifungal resistance.
• The findings pave the way for identifying novel targets for antifungal therapeutics against Candida infections.