YY1 and AP-2 transcription factors interact and cooperate in stimulating ERBB2 gene transcription.

Abstract

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ERBB2 gene is amplified and overexpressed in 30% of human breast cancer. We and other have shown that the overexpression is due to increased transcription rates. AP-2 transcription factors contribute to ERBB2 gene overexpression in breast cancer cells. Our goal is to identify the AP-2 interacting factors contributing to ERBB2 gene overexpression in breast cancer cells. YY1 and AP-2 were shown to interact on the histone H3.2 gene promoter (Wu and Lee, J Biol Chem 2001 276 28). We investigated YY1 and AP-2 factors interaction in breast cancer cells and the functional significance of this interaction on ERBB2 promoter activity. We detected high levels of YY1 in the breast, liver and colon carcinoma cells we have analyzed. High AP-2 levels were only present in breast cancer cells where ERBB2 was overexpressed. We proved the interaction between the endogenous YY1 and AP-2 transcription factors from breast cancer cells by co-immunoprecipitation. The functional significance of the interaction between YY1 and AP-2 transcription factors was analyzed by co-transfection experiments. Increasing amounts of YY1 expression vector alone were unable to modulate ERBB2 promoter activity. In contrast, AP-2 -alpha, -beta and -gamma expression vectors induced a dose-dependent increase in ERBB2 promoter activity in cells where AP-2 was scarce or absent. Co-transfection of a constant amount of AP-2 expression vector with increasing amounts of YY1 expression vector induced a YY1-dose dependent increase in ERBB2 promoter activity. The analysis of the promoter by gel-retardation experiments did not reveal YY1 binding sites, indicating that YY1 modulates AP-2 transcriptional activity by protein-protein interactions. A variant YY1 deleted of its COOH -terminal DNA binding domain did not stimulate AP-2 induced activation of transcription directed by the ERBB2 promoter. The C-terminal deleted protein did interact with AP-2, as shown by co-immunoprecipitation. Finally, YY1 down-regulation by an anti-sens RNA in BT-474 cells, expressing high levels of both YY1 and AP-2, inhibited transcription from AP-2 responsive ERBB2 promoter fragments. Our results show for the first time the contribution of YY1 transcription factor to AP-2 induced stimulation of ERBB2 gene expression in breast cancer cells. YY1 seems to act by interacting with AP-2 and allowing the recruitment of additional factors, probably factors that recognize the C terminus of the protein. Our work contributes to a better understanding of ERBB2 gene overexpression in breast cancer cells.