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Fig. 8. Expression of individual mammalian PKC isoforms or of Bcl-xL in S. cerevisiae is not affected by co-expression of both proteins. (A) Comparable levels of PKC isoform expression were detected in protein extracts obtained from yeast expressing PKC-, -, - or - (PKC-pOW4 lanes) and from yeast co-expressing the PKC isoform and Bcl-xL (PKC-Bcl-xL lanes), after incubation in 2% (w/v) galactose selective medium. Protein extracts obtained from yeast co-transformed with the empty vectors were used as negative controls (YEp51-pOW4 or YEplac181-pOW4 lanes). (B) Comparable levels of Bcl-xL expression were detected in protein extracts obtained from yeast expressing Bcl-xL (YEp51-Bcl-xL or YEplac181-Bcl-xL lanes) and from yeast co-expressing the PKC isoform and Bcl-xL (PKC-, -, - or --Bcl-xL lanes), after incubation in 2% (w/v) galactose selective medium. Protein extracts obtained from yeast co-transformed with the empty vectors were used as negative controls (YEp51/pOW4 or YEplac181-pOW4 lanes). Protein extracts (10-15 µg/lane) were separated on 15% SDS-PAGE followed by western blot analysis. For PKC isoform detection, a specific rabbit antibody to each PKC isoform, followed by horseradish peroxidase-conjugated goat anti-rabbit IgG, was used. For Bcl-xL detection, anti-Bcl-xL goat polyclonal antibody, followed by horseradish peroxidase-conjugated rabbit anti-goat IgG, was used. For ß-actin detection, used as a loading control, membranes were stripped and then reprobed with an anti-actin rabbit antibody. Immunoblots were developed by enhanced chemiluminescence

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