FIG. 3.

Heterologous expression in S. cerevisiae erg4. The yeast erg4 mutant was transformed with V5/6×His-tagged or GFP-tagged Erg4-, DHCR24-, or CBU1206-encoding constructs. Fusion protein expression was induced as described in Materials and Methods. (A) Immunoblotting with antibody directed against the V5 tag showed induced expression of Erg4, DHCR24, and CBU1206. Alpha-tubulin was probed as a loading control. (B) Heterologous expression of CBU1206 and the human Δ24 sterol reductase DHCR24 did not affect growth of a yeast erg4 mutant. The OD₆₀₀ of cultures was measured at the indicated time points following galactose induction. (C) CBU1206-GFP localizes to the yeast ER. The yeast erg4 mutant expressing GFP fusion proteins was fixed and visualized by fluorescence microscopy. As previously described for Erg4 (27), DHCR24 and CBU1206 localized to the ER. GFP (vector control) was cytoplasmic.