Effect of Deptor KD on cell cycle in C2C12 myoblasts. Myoblasts were transfected with either control (scramble) shRNA or shRNA targeting Deptor. Myoblasts were grown in DMEM supplemented with 10% FBS for 18–24 h and stained with propidium iodide to assess cell cycle using FACS. Representative forward scatter histograms highlighting G1 and G2 phases of cell cycle for scramble control (A) and KD (B) are shown. The percentage of cells in each stage of the cell cycle for each treatment group is shown in the accompanying pie graphs. FL2-A, propidium iodide fluorescence. (C) Western blotting of samples (normalized to total protein content for loading) as described above. Blots were probed with total Deptor antibody to show the different groups (top band) and with phospho-specific antibody to detect phosphorylation on S807/S811 of pRb protein (middle band). The lower band shows β-tubulin to confirm equal loading. (D) Quantification of Western blot from C for phosphorylation on S807/S811 of the pRb protein.