Figure 5. Effect of overexpression of nine different PKC isoforms on UGT1A6 protein levels in HEK293T cells.

A) Immunoblot of UGT1A6 coexpressed with the catalytic domains of different PKC isoforms (α, β1, β2, δ, ε, γ, ι, η, and ζ) and β-galactosidase (as a transfection control) in HEK293T cells. Instead of PKC, some cells were cotransfected with the PKC vector control (labeled UGT1A6 control) or with a non-specific protein control (labeled TMED7). Cell lysates were prepared 48 hours post-transfection and subjected to immunoblot analysis with an anti-UGT1A peptide antibody. A UGT1A6 standard (6 to 100 ng per lane) provided with the WB-UGT1A6 kit (BD Gentest) was used as a standard for quantitation purposes. Shown is a representative blot from 3 independent experiments. B) Quantification of UGT1A6 protein expression by immunoblot analysis (representative blot shown in A). Data points correspond to the mean ± standard error of three independent experiments performed on separate days. C) UGT1A6 protein data (from B) normalized to β-galactosidase activities (shown in Figure 4B) Data points on B and C correspond to the mean ± SE of three independent experiments performed on separate days. Statistical comparisons were made using one-way ANOVA followed by a Dunnett’s post hoc test. * indicates that the UGT1A6 protein level is significantly different (P<0.05) from the UGT1A6 control value.