Validation of the Saccharomyces cerevisiae ace1Δ Mutant as a Sensitive Platform for the Identification of Metallophores

Copper is essential for cellular metabolism but becomes highly toxic when intracellular homeostasis is disrupted, promoting reactive oxygen species (ROS) generation and oxidative stress. Here, we established the Saccharomyces cerevisiae ace1Δ mutant as a sensitive biological platform for screening synthetic metallophores that modulate intracellular copper availability. Wild-type BY4741 and ace1Δ strains were exposed to CuSO₄, and cell survival, ROS production, and the expression of copper homeostasis genes were evaluated. The ace1Δ mutant displayed markedly greater copper sensitivity, increased ROS accumulation, and an impaired transcriptional response compared with the wild-type strain. Pre-incubation with N-acylhydrazonic compounds significantly improved survival, with X1INH, X1Fur, and HPCIH providing the strongest protection. Notably, these compounds also rescued the highly copper-sensitive ace1Δ mutant, demonstrating their ability to mitigate copper-induced toxicity. These findings highlight the essential role of Ace1 in copper tolerance and establish the ace1Δ strain as a robust platform for identifying metallophores with potential therapeutic applications.