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. Author manuscript; available in PMC: 2012 Oct 28.
Published in final edited form as: Cell. 2011 Oct 28;147(3):565–576. doi: 10.1016/j.cell.2011.08.050

Figure 7. Myod1 expression redirects Smad3 occupancy in mES cells.

Figure 7

(A) Experimental model. Smad3 co-occupies the genome with Oct4 in mES cells. Myod1 expression was induced in mES cells for 5 days in standard mES cell culture conditions. ChIP-seq was performed to determine if Smad3 was directed to new sites occupied by Myod1.

(B) Oct4 expression is maintained despite induction of Myod1. mES cells containing dox-repressible Myod1 (Nishiyama et al., 2009) were cultured in standard mES cell conditions for 5 days with and without dox. Western analysis was performed to detect expression of Myod1 (top) and Oct4 (middle). TBP was used as a loading control (bottom).

(C) Smad3 occupies new sites with Myod1. The percentage of Smad3 sites in mES cells that are also occupied by Myod1 in myotubes (y-axis) is shown for mES cells without induction of Myod1 (No Myod1) and with induction of Myod1 (+Myod1). The 1000 strongest Smad3 binding sites in each condition were used for this calculation.

(D) Smad3 continues to occupy sites bound by Oct4. Gene tracks show binding of Oct4, Smad3 without induction of Myod1 (red), Smad3 with induction of Myod1 (brown) and Myod1 (after induction) in mES cells at Lefty2 and Tdgf1. Myod1 binding in myotubes is shown at the bottom. The floor is set at 3 counts.

(E). Smad3 occupies new sites bound by Myod1. Gene tracks show binding of Oct4, Smad3 without induction of Myod1 (red), Smad3 with induction of Myod1 (brown) and Myod1 (after induction) in mES cells at the muscle specific genes Mef2d and Ckm. Myod1 binding in myotubes is shown at the bottom.