Figure 2. REST abundance is dependent on the amount of cellular USP15. (A) Multiple siRNAs for USP15 but not USP4 or USP11 reduce the amount of REST. siRNA-mediated knockdown was repeated with five individual USP15 oligos and compared with pooled siRNAs targeting USP4 and USP11. After 72 h, proteins were prepared from subcellular fractions and immunoblotted. Representative blots for REST in the nucleoplasmic fraction are shown with mean quantification normalized to actin and the appropriate siRNA control below (n = 3 individual experiments, error bars show standard deviation). (B) Confirmation of specificity for siRNAs targeting the USP15 paralogs. Cytoplasmic protein extracts from the experiments shown in (A) were immunoblotted for USP15 (polyclonal, *indicates cross-reactivity with USP11), USP4 and USP11 to evaluate knockdown efficiency. (C) The amount of REST correlates with USP15 levels. A scatter plot representing the mean quantification of REST (x-axis) plotted against that of USP15 (y-axis) for each of the controls or samples treated with siRNA targeting USP15, USP4 or USP11 in these experiments; linear regression R² = 0.69.