Selective deletion of enhancer binding sites can change the steady state response characteristics.
A. Deletion of any of the enhancer binding sites from the Scl+19 or Gata2-3 enhancers eliminates the high expression state of Scl, Gata2 and Fli1 seen in the wildtype HSCs. Black crosses mark the deleted sites, red crosses mark the interactions that are no longer significant as a result of the deletion. B. Mutations in the Scl or Fli1 binding site in the Fli1+12 enhancer allow triad activation but lead to reversible bistability-the ON state switches back to OFF in the absence of Notch and Bmp4. C. Deletion of the primary Gata2 binding site from the Fli1+12 enhancer makes the Scl interaction with the enhancer insignificant. This effectively makes Fli1 independent of external regulators Scl and Gata2. Fli1 expression is low for these mutants and monostable. Notch has no effect on Fli1 concentration. Gata2 and Scl show reversible bistability in response to Notch and Bmp4 in these mutants.