Figure 7.

Overexpression of MMS21 suppresses the temperature sensitivity of smc5 cells. (a) SMC5 (yGC141), smc5-31 (yGC231), and smc5-33 (yGC233) cells were transformed with either pRS316 or a 2μ vector with the indicated gene driven by the GAL1 promoter. Fifteen thousand, 3000, 600, 120, and 24 cells were plated on prewarmed SC–ura–leu medium containing 2% dextrose or 2% galactose as indicated. Cells were allowed to grow for 3.5 days. Equal growth was seen on dextrose at 36° (data not shown). Transformants of strains with an intact chromosomal SMC5 locus performed identically to yGC141 transformants under all conditions in these assays (data not shown). Three transformants per strain were tested and the experiment was performed twice. Overexpression of the relevant protein was assayed by Coomassie stain (data not shown). (b) Mms21p suppression of smc5 in liquid culture. smc5-31 and smc5-33 strains with either pRS316 or GAL1-inducible MMS21 were grown in dextrose until midlog and then shifted to 36° SC–ura Gal medium. The doubling times of smc5-31 + Mms21p and smc5-33 + Mms21p strains are 16.3 and 17.7 hr, respectively. (c) Plates from a were grown at 22° for 3 days following 3.5 days of growth at 36°. (d) Overexpression of Nse4p causes DNA damage hypersensitivity. This was done as in a, except dilutions were spotted on SC–ura–leu plates with 25 mm hydroxyurea.