Transcription Factor AP-2 Regulatory Signatures in Breast Cancer

Abstract

AP-2 transcription factors are highly conserved basic helix-span-helix proteins whose members ((x, ß, y, S and c) are crucial regulators of bryonic development. They also play an important role in human neoplasia. uohis ochemical studies have detected high levels of AP-2y expression in primary tumo of breast cancer patients. This high expression has been correlated with reduced survival in all patients and reduced survival in an ERa positive subset treated with hormone therapy. In breast cancer cell lines, AP- 2 factors have been implicated in the regulation of the ERBB2 proto-oncogene and ERa. In an effort to further understand the role of AP-2y in breast carcinoma, this study has sought to identify additional AP-2 activated cellular pathways and ultimately novel transcriptional targets for AP-2 through the use of gene expression profiling. RNAi using three independent AP-2y targeting sequences, has been used to deplete AP- 2y levels in the ERa positive MCF-7 breast carcinoma cell line, chosen as it exclusively expresses the AP-2y family member. Microarrays were then utilised to create an AP-2y dependent transcription profile. Statistical comparisons between non-silencing control siRNA and AP-2y targeting siRNA groups identified a total of 162 gene expression changes (p<0.01). These changes implicate AP-2y in the control of cell cycle progression and developmental signalling. Indeed a role for AP-2y in the control of cell cycle, in particular at the GUS transition, has been verified using flow cytometry. Several of these gene expression changes, including IGFBP3, Transgelin and KIAA1324, have been confirmed using qPCR and immunoblotting. Finally, elevated levels of p21 mRNA and protein have been observed following AP-2y silencing in MCF-7 cells. Additionally, the activity of a p21 promoter reporter is repressed following transfection with an AP-2y expression construct in HepG2 cells. These results coupled with ChIP experiments showing AP-2y occupancy at the proximal promoter region of p21 in cycling MCF-7 cells, implicate AP-2y in the repression of p21 transcription and suggest a role for AP2y in- the, control of cell cycle in breast carcinoma in part through the transcriptional repression of p21.