FOXA1 is an essential determinant of ER expression and mammary ductal morphogenesis

Department of Pharmacology, Case Western Reserve University School of Medicine, 10900 Euclid Avenue, Cleveland, OH 44106, USA.
Development (Impact Factor: 6.46). 06/2010; 137(12):2045-54. DOI: 10.1242/dev.043299
Source: PubMed


FOXA1, estrogen receptor alpha (ERalpha) and GATA3 independently predict favorable outcome in breast cancer patients, and their expression correlates with a differentiated, luminal tumor subtype. As transcription factors, each functions in the morphogenesis of various organs, with ERalpha and GATA3 being established regulators of mammary gland development. Interdependency between these three factors in breast cancer and normal mammary development has been suggested, but the specific role for FOXA1 is not known. Herein, we report that Foxa1 deficiency causes a defect in hormone-induced mammary ductal invasion associated with a loss of terminal end bud formation and ERalpha expression. By contrast, Foxa1 null glands maintain GATA3 expression. Unlike ERalpha and GATA3 deficiency, Foxa1 null glands form milk-producing alveoli, indicating that the defect is restricted to expansion of the ductal epithelium, further emphasizing the novel role for FOXA1 in mammary morphogenesis. Using breast cancer cell lines, we also demonstrate that FOXA1 regulates ERalpha expression, but not GATA3. These data reveal that FOXA1 is necessary for hormonal responsiveness in the developing mammary gland and ERalpha-positive breast cancers, at least in part, through its control of ERalpha expression.

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Available from: Gina M Sizemore, Sep 06, 2014
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    • "In contrast, both wild type and truncated GATA3 in MCF7 were only moderately affected by hormone. FOXA1, an essential determinant of ERα expression [30] and a frequent binding partner of ERα and GATA3 [17,31], decreased in abundance following estradiol treatment in T47D, although not to the same extent as GATA3 and ERα (Figure 4A, B). In MCF7, hormone had little to no impact on FOXA1 levels. "
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    ABSTRACT: The transcription factor GATA3 is a favorable prognostic indicator in estrogen receptor-alpha (ERalpha)-positive breast tumors in which it participates with ERalpha and FOXA1 in a complex transcriptional regulatory program driving tumor growth. GATA3 mutations are frequent in breast cancer and have been classified as driver mutations. To elucidate the contribution(s) of GATA3 alterations to cancer, we studied two breast cancer cell lines, MCF7, which carries a heterozygous frameshift mutation in the second zinc finger of GATA3, and T47D, wild-type at this locus. Immunofluorescence staining and subcellular fractionation were employed to verify cellular localization of GATA3 in T47D and MCF7 cells. To test protein stability, cells were treated with translation inhibitor, cycloheximide or proteasome inhibitor, MG132, and GATA3 abundance was measured over time using immunoblot. GATA3 turn-over in response to hormone was determined by treating the cells with estradiol or ERalpha agonist, ICI 182,780. DNA binding ability of recombinant GATA3 was evaluated using electrophoretic mobility shift assay and heparin chromatography. Genomic location of GATA3 in MCF7 and T47D cells was assessed by chromatin immunoprecipitation coupled with next-generation sequencing (ChIP-seq). GATA3 localized in the nucleus in T47D and MCF7 cells, regardless of the mutation status. The truncated protein in MCF7 had impaired interaction with chromatin and was easily released from the nucleus. Recombinant mutant GATA3 was able to bind DNA to a lesser degree than the wild-type protein. Heterozygosity for the truncating mutation conferred protection from regulated turnover of GATA3, ERalpha and FOXA1 following estrogen stimulation in MCF7 cells. Thus, mutant GATA3 uncoupled protein-level regulation of master regulatory transcription factors from hormone action. Consistent with increased protein stability, ChIP-seq profiling identified greater genome-wide accumulation of GATA3 in MCF7 cells bearing the mutation, albeit with a similar distribution across the genome, comparing to T47D cells. We propose that this specific, cancer-derived mutation in GATA3 deregulates physiologic protein turnover, stabilizes GATA3 binding across the genome and modulates the response of breast cancer cells to estrogen signaling.
    Full-text · Article · Apr 2014 · BMC Cancer
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    • "FOX proteins, specifically the FOXA subclass, have attracted considerable attention for their capacity to impact local chromatin architecture, their essential role in priming for temporal patterns of gene expression observed in early organogenesis and their influence over tissue-specific patterns of NR target gene expression, of particular interest in hormone-related cancers (9,10,18,19). In particular, FoxA1, expressed alongside ER and AR in the developing and mature mammary and prostate ductal epithelia, respectively (20–23), has been shown to contribute significantly to maintaining the oncogenic functions of these nuclear receptors in both treatment-sensitive and -resistant breast and prostate cancers (11,16). While the role of FoxA1 in ER-positive breast cancers appears to be the positive regulation of ER-chromatin binding, resulting in canonical and noncanonical ligand-dependent ER target gene expression and breast cancer cell proliferation (11,24), a more complex relationship between AR and FoxA1 exists in prostate cancer. "
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    ABSTRACT: In prostate cancer, androgen receptor (AR) binding and androgen-responsive gene expression are defined by hormone-independent binding patterns of the pioneer factors FoxA1 and GATA2. Insufficient evidence of the mechanisms by which GATA2 contributes to this process precludes complete understanding of a key determinant of tissue-specific AR activity. Our observations suggest that GATA2 facilitates androgen-responsive gene expression by three distinct modes of action. By occupying novel binding sites within the AR gene locus, GATA2 positively regulates AR expression before and after androgen stimulation. Additionally, GATA2 engages AR target gene enhancers prior to hormone stimulation, producing an active and accessible chromatin environment via recruitment of the histone acetyltransferase p300. Finally, GATA2 functions in establishing and/or sustaining basal locus looping by recruiting the Mediator subunit MED1 in the absence of androgen. These mechanisms may contribute to the generally positive role of GATA2 in defining AR genome-wide binding patterns that determine androgen-responsive gene expression profiles. We also find that GATA2 and FoxA1 exhibit both independent and codependent co-occupancy of AR target gene enhancers. Identifying these determinants of AR transcriptional activity may provide a foundation for the development of future prostate cancer therapeutics that target pioneer factor function.
    Full-text · Article · Jan 2014 · Nucleic Acids Research
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    • "A possible explanation for the discrepancy could be due to differences in estrogen conditions in the experimental designs. Studies investigating the links between ERα, GATA3, and FOXA1 demonstrate that these factors are involved in complex cross-regulatory loops [32], [33]. GATA3 regulates the expression of both ESR1 and FOXA1 mRNA, while ERα regulates GATA3 mRNA. "
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    ABSTRACT: Expression of the estrogen receptor-α (ERα) gene, ESR1, is a clinical biomarker used to predict therapeutic outcome of breast cancer. Hence, there is significant interest in understanding the mechanisms regulating ESR1 gene expression. Proteasome activity is increased in cancer and we previously showed that proteasome inhibition leads to loss of ESR1 gene expression in breast cancer cells. Expression of ESR1 mRNA in breast cancer cells is controlled predominantly through a proximal promoter within ∼400 base pair (bp) of the transcription start site (TSS). Here, we show that loss of ESR1 gene expression induced by the proteasome inhibitor bortezomib is associated with inactivation of a distal enhancer located 150 kilobases (kb) from the TSS. Chromatin immunoprecipitation assays reveal several bortezomib-induced changes at the distal site including decreased occupancy of three critical transcription factors, GATA3, FOXA1, and AP2γ. Bortezomib treatment also resulted in decreased histone H3 and H4 acetylation and decreased occupancy of histone acetyltransferase, p300. These data suggest a mechanism to explain proteasome inhibitor-induced loss of ESR1 mRNA expression that highlights the importance of the chromatin environment at the -150 kb distal enhancer in regulation of basal expression of ESR1 in breast cancer cells.
    Full-text · Article · Dec 2013 · PLoS ONE
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