A novel circuit overrides Adr 1p control during expression of Saccharomyces cerevisiae 2-trans-enoyl-ACP reductase Etr 1p of mitochondrial type 2 fatty acid synthase

摘要

The significance of the chronicled role of the yeast transcription factor Adr1p in regulating ETR1 was examined in wild type and isogenic adr1Δ mutant cells. An ETR1-lacZ reporter construct was used to verify Adr1p-dependent gene expression. On solid glycerol medium containing X-gal, wild-type cells expressing the reporter turned blue, whereas the adr1Δ mutants remained white. β-Galactosidase activity measurements following 24-h cell growth in liquid glycerol medium revealed a 6.5-fold greater expression level of the reporter gene in the wild type compared with the adr1Δ mutant. In contrast, immunoblotting showed that Etr1p abundance was essentially indistinguishable between the two strains whereas Cta1p, whose expression depends on Adr1p, was present in the wild-type cells, but not in the mutants. Moreover, enzyme assays conducted on transformed wild-type and adr1Δ mutant cells expressing a plasmid-borne ETR1 tethered behind the native promoter revealed similar levels of reductase activity, and the lipoic acid content in the two parental strains was equivalent. Hence, while Adr1p influenced the transcription levels of ETR1, it did not alter the abundance of Etr1p, the level of reductase activity, or the cellular amount of lipoic acid. The results point toward a potentially novel layer of control for maintaining physiological levels of lipoic acid.