Characterization of Tti2 mutations. (A) Yeast strains CY6872 (lanes 1, 2, and 3), CY6070 (lanes 5, 6, and 7), CY6874 (lanes 9, 10, and 11), and BY4742 (lane 13), were grown to stationary phase in minimal medium, diluted 1:20 in YPD and grown for 8 hr at 30°. Protein was harvested by grinding with glass beads, and the indicated amount of lysate separated by SDS-PAGE (8%), and Western blotted with anti-Myc antibody. (B) CY6070 was transformed with plasmids containing full length TTI2, or derivatives with C-terminal deletions to residues 350 (Δ351–421) or 325 (Δ326–421). Individual transformants were grown in YPD and spotted serially on a minimal plate containing 5-FOA. (C) Strains CY6070 (DED1-TTI2) and CY6971 (GAL10-TTI2) were grown to stationary phase, then spotted in 10-fold serial dilutions onto YP plates containing 2% galactose, raffinose, or glucose, and grown for 3 d at 30°.