Mecanismos moleculares de regulação da homeostase de cobre em S. cerevisiae: papel das proteínas de detoxificação

Abstract

Copper (Cu) homeostasis is vital for cellular health, involving a complex network of proteins that regulate the transport and detoxification of this metal. Key proteins include the transporters Ctr1 and Fet4, the metallothioneins Cup1 and Crs5, Cu-buffering proteins such as Sod1, and the transcription factors Ace1 and Mac1, which control the expression of genes in response (Cu detoxification and import) to high and low Cu concentrations, respectively. Dysfunction in Cu homeostasis can lead to the accumulation of this metal, resulting in the production of reactive oxygen species, oxidative stress, and loss of cell viability. Investigating the cellular response to high Cu concentrations is essential to understanding cellular adaptations to metal-induced stress. This study aims to investigate the role of different proteins involved in Cu homeostasis in Saccharomyces cerevisiae BY4741 and its isogenic ace1Δ, crs5Δ, and sod1Δ strains exposed to high Cu concentrations. The results showed that BY4741 cell survival was reduced in response to CuSO₄-induced stress, while the ace1Δ strain exhibited greater Cu sensitivity, with a decline in viability starting at 2.5 mM. The crs5Δ and sod1Δ strains displayed increased resistance at high Cu concentrations. Intracellular metal analysis revealed significant variations in wild-type and ace1Δ strains, reflecting the impact of Cu on the homeostasis of other metals. The determination of metallothionein levels indicated that the ace1Δ strain failed to properly regulate the synthesis of these proteins, highlighting the importance of the Ace1 transcription factor. Both the wild-type and ace1Δ strains showed a reduction in total thiols after 15 minutes of Cu exposure, whereas the crs5Δ strain exhibited increased thiol levels after 60 minutes of Cu stress. Intracellular oxidation increased in all tested strains. CTR1 expression decreased after Cu exposure in BY4741 and ace1Δ strains, while CUP1 expression showed a transient increase in BY4741. In the ace1Δ strain, CUP1 expression was not detected. This research emphasizes the critical role of the Ace1 transcription factor and metallothioneins in the response of S. cerevisiae to Cu excess, suggesting that the modulation of Cu homeostasis is essential for cell survival under stress conditions.