Figure 4.

gcd1 expression is regulated by zinc in a Loz1-dependent manner. A, β-galactosidase activity was measured in wild-type and loz1Δ cells expressing the gcd1-lacZ reporter following growth overnight in ZL-EMM or ZL-EMM supplemented with 0, 1, 10, or 200 μm Zn²⁺. Results show average values from 3 independent experiments. B, RNA blot analysis of total RNA purified from wild-type and loz1Δ cells grown overnight in ZL-EMM or ZL-EMM supplemented with 1, 10, or 100 μm Zn²⁺, or C, to exponential phase in YES medium. Blots were probed for the indicated transcripts. Ribosomal RNAs are shown as loading controls. D, immunoblot analysis of crude protein extracted prepped from wild-type cells and cells expressing Gcd1–13×Myc from its endogenous promoter (Gcd1-myc). Immunoblots were probed for c-Myc and loading control Actin (Act1). The positions of the 50- and 37-kDa molecular mass markers are shown on the left. E and F, the indicated strains were grown overnight in ZL-EMM or ZL-EMM supplemented with 200 μm Zn²⁺ and gluconate levels were measured using an enzymatic assay. Data represents the average values from 3 independent experiments with error bars representing standard deviations.