Figure 3.

Overexpression of 444B cures weak [PSI⁺] variants but not strong [PSI⁺] variants. A, effect of overexpression of 444B in different [PSI⁺] variants. Yeast in raffinose medium were grown overnight in galactose medium to induce 444B expression. Yeast were plated on ½ YPD medium before (columns I and III) and after induction of 444B (columns II and IV). Note [psi⁻] yeast in the 779-6A and the 74D-694 backgrounds give different red hues on ½ YPD plates. B, curing of different [PSI⁺] variants was measured in yeast overexpressing 444B. 444B was expressed from the GAL1 promoter in different [PSI⁺] variants and plated at the indicated times. Curing of [PSI⁺] by overexpression of 444B was also done in the 1509 [PSI⁺] ^DM yeast. The 1509 yeast was derived by integrating 444B into the HSP104 chromosomal locus of the 1074 yeast. C, overexpression of 444B causes loss of detectable foci in weak but not strong [PSI⁺] variants. Fluorescence images of GFP-labeled Sup35 are of weak L2888 [PSI⁺] variant overexpressing 444B for one generation and strong SY80 [PSI⁺] variant overexpressing 444B for 15 generations. The L2888 [PSI⁺] yeast were imaged both before stress and after stress. D, rate of curing of different [PSI⁺] variants was determined upon overexpression of 444BΔC19. Curing was measured at the indicated times using the red/white colony assay. E, 5-fluoroorotic acid (5-FOA) shuffle experiment using 1408 yeast in which the pJ312 plasmid, which encodes Sc-Hsp104 under the control of the S. cerevisiae HSP104 promoter on a URA3-based centromeric plasmid., was shuffled with plasmids expressing either 444B or 444BΔC19. The 444B and 444BΔC19 were expressed using the Sc-HSP104 promoter. Only constructs that support [PSI⁺] propagation grow on plates without adenine.