Figure 5.

OTUD7B‐dependent regulation of LSD1 fluctuation is crucial for cell cycle progression. A) MDA‐MB‐231 cells stably expressing either OTUD7B sgRNA (top panel) or LSD1 shRNA (bottom panel) were synchronized by thymidine (2 × 10⁻⁶ m) and nocodazole (200 ng mL⁻¹) treatment, released and harvested at the indicated time points. Cell lysates were subjected to IB analysis as indicated. Asy: asynchronous cells. SE: shorter exposure; LE: longer exposure. B) LSD1^2KR‐reconstituting cells infected with sgOTUD7B lentiviruses were synchronized and released as in (A), followed by IB analysis. C) MDA‐MB‐231 cells infected with lentiviral p62 shRNA were synchronized as in (A), released and harvested at the indicated times, followed by IB analysis. D) MDA‐MB‐231 cells transfected with the indicated Ub constructs were synchronized as in (A) and released. Cell lysates were subjected to IP and IB analysis as indicated. E,F) Parental or LSD1‐reconstituting MDA‐MB‐231 cells infected with the indicated lentiviral constructs were synchronized as in (A), released and collected at the indicated time points. Cell cycle profile was obtained by propidium iodide (PI) staining and fluorescent‐activated cell sorting (FACS) analysis. TN: thymidine and nocodazole treatment. Data shown are representative of three independent experiments.