Figure 5.

Role of the TF Pap1 in the recruitment of Atf1 to one subset of stress promoters. A, Pap1 is required for H₂O₂-dependent expression of ctt1 and srx1 but not for gpd1 and hsp9. Cultures of strains 972 (WT), AV18 (Δsty1), MS98 (Δatf1), and AV25 (Δpap1) were treated with 1 mm H₂O₂ for 15 min or left untreated. Total RNA was obtained and analyzed as in Fig. 1D. B, deletion of Pap1 abolishes the activation of srx1 and ctt1 of cells expressing the phospho-mimicking HA-Atf1.10D. YE cultures of strains 972 (WT), EP203.10D (Δatf1 + HA-Atf1.10D), EP303.10D (Δatf1 Δsty1 + HA-Atf1.10D), and PG125.10D (Δatf1 Δpap1 Δsty1 + HA-Atf1.10D) were treated with 1 mm H₂O₂ for 15 min or left untreated. Total RNA was obtained and analyzed as in Fig. 1D. C, the recruitment of Atf1-HA to the ctt1 and srx1 promoters is dependent on Pap1. MM cultures of strains CS51 (Δpap1 atf1-HA), CS51.7D (Δpap1 atf1.7D-HA), and CS51.7M (Δpap1 atf1.7M-HA) were treated with 1 mm H₂O₂ for 5 min or left untreated, and ChIP experiments were performed as in Fig. 3A. D, oxidized/active Pap1 facilitates the H₂O₂-dependent recruitment of Atf1-HA to the ctt1 and srx1 promoters. MM cultures of strains CS62 (Δtrr1 atf1-HA, constitutively expressing oxidized nuclear Pap1) and CS79.C523D (pap1.C523D atf1-HA, expressing a constitutively reduced nuclear Pap1) were left untreated or treated with 1 mm H₂O₂ for 5 min. ChIP experiments were performed as in Fig. 3A. E, Pap1 binds to the ctt1 and srx1 promoters to the same extent in cells expressing or lacking Atf1. MM cultures of strains 972 (WT) and MS98 (Δatf1) were treated with 1 mm H₂O₂ for 5 min or left untreated, and ChIP experiments using anti-Pap1 antibodies were performed as in Fig. 3A.