FIG 1 .

Expression of the mitochondrial ABC transporter Atm1 is induced by Cu in a Cuf1-dependent manner in C. neoformans. (A) Exponentially growing cultures of C. neoformans strains H99 (DTY758), ΔCuf1 (DTY761), and ΔCuf1 with complementing Cuf1 (DTY762) were treated with 1 mM BCS and 1 mM Cu for 3 h. Gene expression was analyzed by qRT-PCR with specific primers for ATM1 and ACT1 (used for data normalization). n = 4 (one-way repeated-measures ANOVA test, P = 0.02). (B) Exponentially growing cultures of C. neoformans strains Cuf1-FLAG (DTY762) and H99 (DTY758) were treated with 1 mM CuSO₄ or 1 mM BCS. ChIP was performed using anti-FLAG antibody, and Cuf1 occupancy was analyzed by qRT-PCR with primers for the promoter regions of the ATM1 and TUB2 genes. C. neoformans H99 was used as a negative control. n = 3 (Student’s t test of paired samples, P = 0.002). (C) Cellular protein extracts were obtained from exponentially growing cultures of C. neoformans strain Atm1-F (DTY947) that had been left untreated (unt) or had been treated with 1 mM Cu during the indicated times and analyzed by SDS-PAGE and immunoblotting with antibodies against FLAG and histone 3 (H3; loading control). (D) S. cerevisiae Atm1 protein expression is not affected by Cu stress. Cellular protein extracts were obtained from untreated or Cu (1.25 mM)-treated exponentially growing cultures of S. cerevisiae strain W303A (wild type [WT]) and analyzed by SDS-PAGE and immunoblotting with antibodies against Atm1 and porin (loading control).