Isw2-mediated chromatin remodeling governs antifungal tolerance and heteroresistance in Candida albicans

Antifungal tolerance, unlike resistance, allows cells to grow slowly at concentrations above the minimum inhibitory concentration. While resistance mechanisms are well characterized, the pathways underlying tolerance remain elusive. Here, we performed a genetic screen of a transcriptional factor mutant library to identify regulators of azole tolerance in Candida albicans. This screen uncovered Isw2, the catalytic subunit of the ISW2 ATP-dependent chromatin remodeling complex, as a negative regulator of azole tolerance. Integrating transcriptomics, Isw2 DNA-binding profiles, and nucleosome-occupancy analyses revealed that Isw2 maintains a repressive chromatin architecture at the CRZ1 promoter, limiting the nucleosome-depleted region and suppressing CRZ1 transcription. Isw2 also modulates fluconazole heteroresistance and amphotericin B sensitivity through Crz1. In addition, we identified the copper-responsive transcription factor Mac1 as a context-dependent regulator of azole tolerance, acting negatively under copper limitation but positively under copper-replete conditions. Together, these findings reveal unexpected roles for chromatin remodeling and copper homeostasis in antifungal tolerance.