Figure 4.

Depletion of Gata3 in luminal tumor cells promotes basal-like differentiation in vitro. (A, B) Luminal mammary tumor cells from MMTV-PyMT mice were infected with psi-LVRU6GP-control (sh-Ctrl) or psi-LVRU6GP-Gata3 targeting different sequences of mouse Gata3 (sh-Gata3-a and sh-Gata3-c), selected with puromycin, and analyzed by phase-contrast and fluorescence microscope (A) or western blot (B). Note that sh-Ctrl cells exhibited cobblestone morphology and close-contact with neighboring cells at cell junctions whereas sh-Gata3 cells displayed an elongated and spiky appearance and were isolated. (C, D) MMTV-PyMT luminal tumor cells infected with sh-Ctrl and sh-Gata3-c were analyzed by qRT-PCR. Data represent the mean ± SD from triplicates of primary tumor cells of each group. The asterisk (*) denotes a statistical significance from sh-Ctrl and sh-Gata3-c samples determined by student's t-test. (E) Representative immunofluorescent staining analysis of the MMTV-PyMT luminal tumor cells infected with sh-Ctrl and sh-Gata3-c. Cells were immunostained with antibodies against eGFP (green) and Ck14 (red). Percentage of Ck14 positive cells in the eGFP positive cell population was calculated. Data represent the mean ± SD from more than 500 eGFP positive cells in five randomly selected fields for each group of the two independent experiments.