Yeast strains and plasmids are listed in tables S1 and S2, respectively. Standard techniques were used for strain construction and plasmid transformation. Cells were cultured at 30°C. YPD medium consisted of 1% yeast extract, 2% Bacto peptone, and 2% dextrose. YPGal medium contained 2% galactose instead of dextrose. Plasmid selection was by either supplementation of rich media with ClonNAT (100 μg/ml) or omission of uridine from synthetic media, as appropriate. Note that C-terminal 3×HA tagging of Yap8 did not support wild-type arsenic resistance; therefore, an N-terminal tag was used for immunoblot analysis and fluorescence microscopy. Human cell lines (U937 and Ramos) were cultured in suspension under standard conditions.

pJH62 is a low-copy centromeric plasmid derived from ycPlac22 (38) in which the TRP1 marker has been replaced with the NAT-MX6 cassette, allowing for selection by ClonNAT resistance (39). Site-directed mutagenesis was performed using the QuikChange method (Agilent). Mutations were verified by sequencing.

Sodium arsenite (Sigma-Aldrich) was used at 1 mM or less, a standard concentration in yeast (1718, 21, 33). We previously showed that cells retained viability under these conditions for up to (at least) 4 hours (13). Within just 1 to 2 hours after drug washout, the cells resumed growth with a normal doubling time, indicating that the cellular responses observed during treatment are likely to be physiologically relevant and specific to arsenic toxicity, rather than reflecting nonspecific toxicity in dead or dying cells (13).

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