Figure 2. Identification of mediators of doxorubicin induced apoptosis.

To determine which genes identified by our screen were true or false positives, we targeted each for degradation by shRNA. (A) Individual genes listed in 1B were targeted for shRNA mediated degradation in U2OS cells. shRNA targeted and control cells were treated with 400 ng/ml doxorubicin and measured by propidium iodide (PI) assay 72 hours later. Levels of apoptosis are reported as % apoptosis in shRNA targeted cell compared with vector control cells. Five of the cell lines appeared to be false positives and did not display reduced doxorubicin induced apoptosis. The other lines were impaired by 20–40% for doxorubicin induced apoptosis. (B) Knockdown levels in these cell lines were determined by qPCR comparing with vector control cells and listed as % remaining expression in target cells in 2A. Genes are listed in the order presented in 2B.