Figure 1

Establishment of the conditional ΔAP-2γ expression system using the Tet-ON ^® System in N202.1A breast cancer cells. (A) The schematic representation of the structural AP-2 protein domains shows the N'-terminal proline- and glutamine-rich (PY) transactivation domain, the DNA-binding domain which consists of the basic region and the helix-span-helix motif and the C'-terminal dimerization domain. (B) The dominant-negative AP-2γ mutant (ΔAP-2γ) has a deleted transactivation domain and therefore an abolished transactivation potential. It still dimerizes with full length AP-2 proteins, thereby inhibiting their function. (C, D) Usage of a bidirectional Tet Responsive Element promoter (TRE) allowed for conditional coexpression of ΔAP-2γ and EGFP upon addition of doxycycline (+dox, 2 μg/ml) in rtTA containing N202.1A cells. (E-H) Generation of N202.1A clones expressing ΔAP-2γ and/or EGFP respectively: stably transfected N202.1A rtTA breast cancer cells were screened for conditional expression of rtTA and either ΔAP-2γ and EGFP (E, Δ#7) or EGFP only (F, Co#11) using fluorescence (E, F) and phase contrast (G, H) microscopy. Clones display a low backround expression in the uninduced state (-dox) but high transgene expression upon induction (+dox, 2 μg/ml). Scale Bar in E-H represents 50 μm.