Figure 4: RUNX1 prevents oestrogen-mediated AXIN1 repression.

(a) MCF7/shRx1_RUNT^dox cells were maintained in 10% charcoal-stripped serum for 48 h, treated as indicated for the following 48 h, and AXIN1 mRNA levels were measured by RT–qPCR and corrected for 18S RNA (mean±s.e.m. of three independent experiments). (b) MCF7/shRx1_RUNT^dox cells in 10% complete serum were treated as indicated for 48 h, and AXIN1 mRNA levels were measured by RT–qPCR and corrected for 18S RNA (mean±s.e.m. of three independent experiments). *P<0.05 by t-test. (c) Scatter plot of the global E2 responsiveness in the presence (y axis) versus absence (x axis) of RUNX1 in MCF7 cells. (d) The indicated ER⁺ (left) and ER⁻ (right) mammary epithelial cell lines were engineered with the dox-inducible shRx1_3′-UTR lentiviral vector and treated with dox for 4 days before western blot analysis of the indicated proteins. (e) RT–qPCR results for Axin1 and Runx1 from predominantly ER⁺ mature luminal (ML) mammary epithelial cells (left) and predominantly ER⁻ luminal progenitor (LP) cells (right) isolated from RUNX1-knockout and control mammary glands as described in the ‘Methods’ section.