FIG. 4.

(A) Western blot analysis of 1a and 2a protein accumulation in wt and ole1w_i′ yeast containing a plasmid expressing B3CAT and either ADH1-promoted 1a and 2a expression plasmids (lanes 1 to 3) or GAL1-promoted 1a and 2a expression plasmids (lanes 4 to 6). Yeast was grown to mid-log phase in Gal medium containing no UFA (−) or 0.2 mM UFA (+). Cell lysates were prepared, and equal amounts of total protein were electrophoresed and Western blotted as described in Materials and Methods. (B) BMV-directed CAT expression in the yeast cells described in the legend to panel A. (C) Distribution of 1a and 2a between membrane and soluble cytoplasmic fractions in ole1w_i′ yeast with or without the UFA supplementation. ole1w_i′ yeast cells expressing GAL1-promoted 1a, 2a, and RNA3 were harvested at mid-log phase, and total protein (Tot.) was extracted. A portion of the lysate was centrifuged at 10,000 × g to yield pellet (Pel.) and supernatant (Sup.) fractions. The same percentage of each fraction was analyzed by electrophoresis and Western blotting.