Fig. 2.

mph1Δ rescues several defects of smc6 and mms21 mutants, whereas Mph1 overexpression confers opposite effects. (A and B) mph1Δ rescues the growth defects and DNA damage sensitivity of smc6–56 (A), smc6-P4, and mms21–11 (B). WT and mutant cell cultures were diluted and spotted onto plates containing no drug or the indicated concentration of drugs or were treated with the indicated dose of UV light. (C and D) mph1Δ rescues centromere separation defects in smc6–56 cells. Cells were arrested in G1 at 23 °C and were shifted to 37 °C for 1 h to inactivate the smc6–56 allele before being released from G1. Samples were collected every 15 min to examine cell-cycle progression by FACS and chromatid separation by the appearance of 2 dots of GFP-LacI bound to tandem LacO repeats near the centromere on chromosome IV. To examine the events in the first cell cycle only, alpha-factor was added back to the cultures 45 min after release. (E and G) Mph1 overexpression sensitizes cells with defective Smc5/6 complexes. Cells containing smc6-P4 or mms21–11 (E) or SMC6-YFP (SMC6-Y, G) were transformed with pGAL-Mph1 or the control vector; cell cultures were diluted and spotted on plates lacking uracil with glucose or galactose and with and without MMS or HU. (F) Smc6-YFP confers normal growth but is synthetic sick with mms21–11. A representative tetrad from diploid strains heterozygous for SMC6-YFP and mms21–11 is shown. The genotype for each spore clone is indicated. (H) mph1Δ suppresses the lethality of mms21Δ and smc6Δ cells. Representative tetrads from diploid strains with the indicated genotypes are shown. The spore clones containing mms21Δ mph1Δ (Left) and smc6Δ mph1Δ (Right) are underlined; those containing mms21Δ (Left) and smc6Δ (Right) are marked with dotted lines, and their genotypes are deduced from sibling spore clones.