Regulation of KiSS-1 Metastasis Suppressor Gene Expression in Breast Cancer Cells by Direct Interaction of Transcription Factors Activator Protein-2α and Specificity Protein-1

Dianne C Mitchell,Maen Abdelrahim,Jinsheng Weng,Lewis J Stafford,Stephen Safe,Menashe Bar-Eli,Mingyao Liu

doi:10.1074/jbc.m506245200

Abstract

KiSS-1 has been shown to function as a tumor metastasis suppressor gene and reduce the number of metastases in different cancers. The expression of KiSS-1 or KiSS1, like other tumor suppressor, is commonly reduced or completely ablated in a variety of cancers via an unknown mechanism. Here we show that the loss of KiSS-1 expression in highly metastatic breast cancer cell lines correlates directly with the expression levels of two transcription factors, activator protein-2alpha (AP-2alpha) and specificity protein 1 (Sp1), which synergistically activate the transcriptional regulation of KiSS-1 in breast cancer cells. Although the KiSS-1 promoter contains multiple AP-2alpha binding elements, AP-2alpha-mediated regulation occurs indirectly through Sp1 sites, as determined by deletion and mutation analysis. Overexpression of AP-2alpha into highly metastatic breast cell lines did not alter KiSS-1 promoter-driven luciferase gene activity. However, co-transfection of AP-2alpha wild-type or the dominant negative form of AP-2 lacking its C-terminal DNA-binding domain, AP-2B, together with Sp1, increased KiSS-1 promoter activity dramatically, suggesting that AP-2alpha regulation of KiSS-1 transcription does not require direct binding to the KiSS-1 promoter. Furthermore, we demonstrated that AP-2alpha directly interacted with Sp1 to form transcription complexes at two tandem Sp1-binding sites of the promoter to activate KiSS-1 transcription. Together, our results indicate that AP-2alpha and Sp1 are strong transcriptional regulators of KiSS-1 and that loss or decreased expression of AP-2alpha in breast cancer may account for the loss of tumor metastasis suppressor KiSS-1 expression and thus increased cancer metastasis.

Highlights

The vast majority of breast cancer deaths results from complications caused by tumor cell metastasis rather than as a consequence of the original tumor growth
Endogenous KiSS-1 Expression in Breast Cancer Cell Lines Correlates with AP-2␣ Expression Levels—We and other laboratories have reported that KiSS-1 expression is lost in highly metastatic breast cancer cells
Sp1 expression was slightly decreased in the metastatic cancer cell lines (Fig. 1A). These data suggest that the expression of KiSS-1 was correlated very well with the expression levels of the two transcription factors, especially AP-2␣, in breast cancer cells

Summary

Introduction

The vast majority of breast cancer deaths results from complications caused by tumor cell metastasis rather than as a consequence of the original tumor growth. Metastasis suppressor genes, which inhibit the spread of cancers to secondary sites, have become the target of mounting clinical and basic cancer research One such gene, KiSS-1 or KiSS1, was originally identified as a metastasis suppressor by microcell-mediated transfer in melanoma lines, by which it was found to reduce tumor cell invasive and migratory properties without affecting their tumorigenicity [2]. C-terminal amidation of the KiSS-1 peptide leads to strong binding with GPCR54, initiating a series of cellular changes, including increased intracellular [Ca2ϩ] and inositol 1,4,5-trisphosphate release, as well as morphological changes, such as up-regulating focal adhesion and stress fiber formation [4, 8, 10, 13] These signaling events inhibit chemotaxis and invasion, reducing the incidence of tumor metastasis [4, 5, 13]. While much has been learned about the possible physiological effects of KiSS-1 expression in cancer cells, the mechanism controlling KiSS-1 transcriptional regulation is still unknown as is the underlying reason for its loss during metastatic progression

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Results

Conclusion