Figure 1.

USP7 depletion results in decreased Chk1 protein levels. (A) U2OS cells were transfected twice with Luciferase (Luc) or different USP7 siRNA oligos. 72 h later, cells were lysed for analysis by western blot with the indicated antibodies. (B) 293T and U2OS cells were treated with PD22077. At the indicated time points, cells were lysed for analysis by western blot with the indicated antibodies. (C) U2OS cells were transfected twice with the indicated siRNA oligos (20 μM). Of Claspin siRNA oligos, lower amounts were used (0.1 or 0.02 μM). Cells were lysed and analyzed as in (A). (D) U2OS cells were transfected twice with the indicated siRNA oligos. 72 h later, cells were lysed for analysis by western blot with the indicated antibodies. Right panel demonstrates USP29 downregulation in representative samples. (E) U2OS cells were transfected with Luc or USP7 siRNA oligos. After 24 h, cells were simultaneously transfected with empty vector (EV) or HA-Claspin and Luc or USP7 siRNA oligos. The next day, cells were lysed and analyzed as in (A). In all panels: the arrow points to Chk1. The asterisk indicates an aspecific band.