Fig. 3. Myc overexpression does not block differentiation but delays the cell cycle exit of neural cells. (A) After 4 days in 2% FCS both Myc-overexpressing and control cells expressed the differentiation markers GFAP and TUJ-1 although the morphology of the Myc NPCs varies. (B) The expression of the NPC markers Bmi-1 and nestin decrease dramatically during differentiation of NPCs. (C) Double staining shows that GFAP-positive Myc-overexpressing cells, with the morphology of differentiated cells, are not also positive for the progenitor marker Bmi-1. (D) Myc delays the cell cycle exit and increases the number of cells in S phase during differentiation-promoting conditions (2% FCS) for 3 days as demonstrated by BrdU labelling. During the first day 10.8% of the control cells (s.d.=7.2, n=5) and 32.8% of the Myc cells (s.d.=11.7, n=5) were still proliferating. During the second day, 2.1% (s.d.=2.1, n=5) of the control cells and 6.5% (s.d.=6.6, n=5) of the Myc cells had not exited the cell cycle. By day 3 almost none of the control cells (0.0%, s.d.=0.14, n=6) but 4.4% of the Myc-overexpressing NPCs (s.d.=3.9, n=4) were still proliferating (Student's t-test *P=0.041).