Knockdown of the E1 enzyme induces cell death in malignant cells. (A) THP1, K562, U937, and OCI-AML5 leukemia cells were infected with an E1 shRNA lentiviral vector or control sequences, and populations of infected cells were selected. Total cellular proteins were isolated and analyzed by SDS-PAGE followed by immunoblotting with the use of anti-E1, anti-ubiquitin, and anti–α-tubulin antibodies. (B) Cells infected with an E1 shRNA lentiviral vector or control sequences were seeded 24 hours after infection into 96-well plates (5 × 103 cells/well) in the presence of puromycin to select for infected cells. Seven days after seeding, cell growth and viability were assessed by the MTS assay. Data represent the mean percentage ± SD of viable cells relative to cells infected with control sequences (n = 3). A representative experiment is shown. (C) K562 leukemia cells were infected with an E1 shRNA lentiviral vector or control sequences, and populations of infected cells were harvested at increasing times after infection. Total cellular proteins were isolated and analyzed by SDS-PAGE followed by immunoblotting with the use of anti-E1 and anti–α-tubulin antibodies. (D) Cells infected with an E1 shRNA lentiviral vector or control sequences were seeded 24 hours after infection into 96-well plates (5 × 103 cells/well) in the presence of puromycin to select for infected cells. At increasing times after seeding, cell growth and viability were assessed by the MTS assay. Data represent the mean percentage ± SD of viable cells relative to cells infected with control sequences (n = 3). A representative experiment is shown.
