Abstract
BackgroundThe transcription factor DEAF-1 has been identified as a high affinity binding partner of the LIM-only protein LMO4 that plays important roles in mammary gland development and breast cancer. Here we investigated the influence of DEAF-1 on human and mouse mammary epithelial cells both in vitro and in vivo and identified a potential target gene.ResultsOverexpression of DEAF-1 in human breast epithelial MCF10A cells enhanced cell proliferation in the mammary acini that develop in 3D cultures. To investigate the effects of Deaf-1 on mammary gland development and oncogenesis, we generated MMTV-Deaf-1 transgenic mice. Increased ductal side-branching was observed in young virgin mammary glands, accompanied by augmented cell proliferation. In addition, the ratio of the progesterone receptor isoforms PRA and PRB, previously implicated in regulating ductal side-branching, was altered. Affymetrix gene profiling studies revealed Rac3 as a potential target gene and quantitative RT-PCR analysis confirmed that Rac3 was upregulated by Deaf-1 in immortalized mouse mammary epithelial cells. Furthermore, MMTV-Deaf-1 transgenic mammary glands were found to have elevated levels of Rac3 mRNA, suggesting that it is a bona fide target.ConclusionWe have demonstrated that overexpression of Deaf-1 enhances the proliferation of human breast epithelial cells in vitro and mouse epithelial cells in vivo. Transgenic mammary glands overexpressing Deaf-1 exhibited a modest side-branching phenotype, accompanied by an increase in the number of BrdU-positive cells and a decrease in the proportion of PRA-expressing cells. Although proliferation was enhanced in Deaf-1 transgenic mice, overexpression of this gene was not sufficient to induce the formation of mammary tumors. In addition, our studies identified Rac3, encoding a small Rho-like GTPase, as a potential target of Deaf-1 in mouse mammary epithelial cells.
Highlights
The transcription factor Deformed epidermal autoregulatory factor-1 (DEAF-1) has been identified as a high affinity binding partner of the LIM-only protein LMO4 that plays important roles in mammary gland development and breast cancer
DEAF-1 enhances the proliferation of human mammary epithelial cells To generate human breast epithelial cells stably overexpressing human DEAF-1 protein, MCF10A cells were initially transfected with a vector containing the mouse Ecotropic Receptor (EcoR) gene to generate MCF10A-EcoR cells
Western blotting of whole cell lysates confirmed that DEAF-1 was overexpressed in DEAF-1-transduced MCF10A-EcoR cells relative to control cells transduced with an empty vector (Fig. 1A)
Summary
The transcription factor DEAF-1 has been identified as a high affinity binding partner of the LIM-only protein LMO4 that plays important roles in mammary gland development and breast cancer. We investigated the influence of DEAF-1 on human and mouse mammary epithelial cells both in vitro and in vivo and identified a potential target gene. Mammary gland development requires the coordinated action of hormones and growth factors [1]. Functional development of the mammary gland is initiated at puberty upon secretion of the ovarian hormones estrogen and progester-. By 8 – 10 weeks of age in the mouse, complex branching and elongation through the stroma results in an extensive network of ducts filling the entire mammary fat pad. The process of involution commences and involves extensive remodeling of the mammary gland to a virginlike state
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