dun1 ixr1 synthetic lethality is rescued by increased RNR activity.

(A) Schematic representation of the Ixr1 protein. HMG boxes are shown in blue and polyQ regions in black. (B) Alignment of homologous HMG proteins close to the conserved S366. (C) Tetrad analysis demonstrating that deletion of SML1 rescues the dun1Δ ixr1-S366F synthetic lethality. The dun1Δ ixr1-S366F strain (TOY544) carries a DUN1-containing plasmid, pK503, which confers a red color; this plasmid is lost in ixr1-S366F dun1Δ sml1Δ colonies (TOY544×TOY588). (D) Tetrad analysis demonstrating that deletion of SML1 rescues the ixr1Δ dun1Δ synthetic lethality (TOY604×TOY527). (E) Tetrad analysis demonstrating that a low-copy RNR1 vector rescues the ixr1Δ dun1Δ synthetic lethality. The diploid strain ixr1/IXR1 dun1/DUN1 (TOY527×TOY603) was transformed with pRS316 and pBJ6 (pRS316-RNR1), sporulated and tetrad analysis was performed. (F) Overexpression of RNR1 rescues the ixr1-S366F dun1Δ synthetic lethality. The pK503 plasmid, containing the DUN1 gene and conferring red color, is lost on YP-Gal medium in the ixr1-S366F dun1Δ strain transformed with pESC-URA-pGAL1-RNR1 plasmid (left panel, sectoring phenotype), but not when transformed with pESC-URA-pGAL1 (right panel). (G) Low-copy RNR1 vector rescues the ixr1-S366F dun1Δ synthetic lethality. The pK503 plasmid, containing the DUN1 gene and conferring red color, is lost in the ixr1-S366F dun1Δ strain transformed with the pBJ6 plasmid (left panel, sectoring phenotype), but not when transformed with pRS316 (right panel). (H) Overexpression of RNR3 rescues the ixr1-S366F dun1Δ synthetic lethality. The pK503 plasmid, containing the DUN1 gene and conferring red color, is lost in the ixr1-S366F dun1Δ strain transformed with pBAD79 plasmid (left panel, sectoring phenotype), but not when transformed with pRS414 (right panel).