Essential role of cyclin B1 and B2 in cell proliferation and survival. (A) Conditional silencing of cyclin B1. HeLa cells were engineered to stably express ^mAIDcyclin B1, tTA, and TIR1, concurrently disrupting the endogenous cyclin B1 with CRISPR-Cas9. Clones of ^mAIDB1^KOB1 cells were isolated and cultured in the presence of Dox and IAA (DI). The cells were harvested at different time points for immunoblotting analysis. Lysates from control HeLa cells were used to compare endogenous cyclin B1 levels. Equal loading of lysates was confirmed by immunoblotting for actin. (B) Simultaneous silencing cyclin B1 and B2. ^mAIDB1^KOB1B2 cells were generated and treated with DI similarly as in A (see Materials and methods). (C) Conditional silencing cyclin B2. ^AIDB2^KOB2 cells were generated and treated with DI similarly as in A (see Materials and methods). (D) Depletion of cyclin B1 and B2 promotes apoptosis. Different cell lines were cultured with or without DI and harvested at the indicated time points for immunoblotting. (E) Depletion of cyclin B promotes mitotic block and apoptosis. The indicated cell lines were cultured with or without DI. At different time points, the cells were fixed and analyzed with flow cytometry. Positions of 2N and 4N DNA content are indicated. (F) Silencing of cyclin B abrogates clonogenic survival. ^mAIDB1^KOB1B2 cells were cultured with or without DI for 2 wk. Colonies were fixed, stained, and quantified. Mean ± SEM from six independent experiments. Mann–Whitney test: **P < 0.01; ns P > 0.05. Source data are available for this figure: SourceData F1.