(A) LF1 cells were engineered using lentivirus vectors to stably express EZH2 cDNA and a 4-OHT-inducible ER:RAS(G12V) protein. Control cell lines were constructed using empty vectors. Cells were treated with 4-OHT (200 nM) or vehicle (ethanol) for the indicated times, and the levels of EZH2 and ER:RAS proteins were determined by immunoblotting.
(B) Proliferation in the experiment shown in (A) was assessed by counting cell numbers (*p < 0.05, **p < 0.01, n = 3).
(C) SA-β-Gal-positive cells were scored after 12 days of 4-OHT treatment in the experiment shown in (A) (**p < 0.01, n = 3).
(D) p16 mRNA expression was determined by real-time qPCR as in (C) (**p < 0.01, n = 3).
(E) Expression of the indicated SASP genes was determined by real-time qPCR as in (C) (*p < 0.05, **p < 0.01, n = 3).
Error bars represent SD. See also Table S1.