FIGURE 8.

Rescue of lipotoxicity phenotype in H1246. Exogenously supplied ALA inhibits the growth of yeast H1246. This inhibition can be rescued by expressing genes encoding TAG-synthesizing enzymes from flax and the rescuing effect is correlated with the ability to synthesize TAG as demonstrated with Nile red assay and TLC analysis. Growth curves of recombinant yeast H1246 cultured in induction medium with 0 (A), 100 μm (B), and 1 mm (C) ALA, were constructed by measuring the optical density (OD) of the cultures at 600 nm. Data are presented as mean ± S.E. (n = 3 or 6). BY4742 (wild-type) and H1246 harboring the empty vector pYES were used as positive and negative controls, respectively. For negative controls, six independent colonies were analyzed and consistently showed delayed growth and low final density even in the absence of free fatty acid. The panels on the right side, from top to bottom, show the growth of 2.5 μl of each 10-fold dilution (1 to 10⁻⁴) of recombinant yeast H1246 spotted onto induction medium plates containing 0, 100 μm, and 1 mm ALA. The negative controls were annotated as “−” in the image. WT, wild-type; D1-1, LuDGAT1-1; DCR1, LuDCR1; D2-3, LuDGAT2-3; P1, LuPDAT1; P2, LuPDAT2; P6, LuPDAT6.