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The BCL9-2 proto-oncogene governs estrogen receptor alpha expression in breast tumorigenesis.

Zatula N, Wiese M, Bunzendahl J, Birchmeier W, Perske C, Bleckmann A, Brembeck FH - Oncotarget (2014)

Bottom Line: BCL9-2 overexpression in vivo delayed the mammary involution and induced alveolar hyperplasia.We found, that primary cell cultures of BCL9-2 breast tumors responded to tamoxifen treatment.Finally, we describe a novel mechanism, how BCL9-2 regulates ER transcription by interaction with Sp1 through the proximal ESR1 gene promoter.

View Article: PubMed Central - PubMed

Affiliation: Tumor Biology and Signal Transduction, Georg-August-University Göttingen, Germany. Dept. of Hematology and Medical Oncology, Georg-August-University Göttingen, Germany.

ABSTRACT
The majority of human breast cancers express estrogen receptor alpha (ER), which is important for therapy with anti-estrogens. Here we describe the role of BCL9-2, a proto-oncogene previously characterized as co-activator of Wnt/ß-catenin signaling, for mammary tumorigenesis in mice and human. ER positive human breast cancers showed overexpression of BCL9-2 and tamoxifen treated patients with high BCL9-2 demonstrated a better survival. BCL9-2 was upregulated during puberty and pregnancy in normal mammary epithelia, but downregulated in the involuted gland. BCL9-2 overexpression in vivo delayed the mammary involution and induced alveolar hyperplasia. Moreover, aged BCL9-2 transgenic mice developed ductal-like mammary tumors with high nuclear ER expression. We found, that primary cell cultures of BCL9-2 breast tumors responded to tamoxifen treatment. Moreover, BCL9-2 regulated the expression of ER and the proliferation of human breast cancer cells independently of ß-catenin. Finally, we describe a novel mechanism, how BCL9-2 regulates ER transcription by interaction with Sp1 through the proximal ESR1 gene promoter. In summary, BCL9-2 induces ER positive breast cancers in vivo, regulates ER expression by a novel ß-catenin independent mechanism in breast cancer cells, and might predict the therapy response to tamoxifen treatment.

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Primary breast tumor cells from BCL9-2 transgenic mice respond to estrogen and tamoxifen(A) Bright field image and co-immunofluorescence stains of primary cells established from mammary tumors of BCL9-2 females. Merged pictures of the co-stains with the indicated antibodies are shown. (B) Dose-response analysis of primary control and of primary BCL9-2 tumor cells for increasing concentrations of estrogen (E2) and tamoxifen (TAM), relative to vehicle treated cells. Cells were stimulated with the indicated concentrations for 48 hours and cell viability was determined by MTT assays. (C-F) Proliferation and colony formation of primary BCL9-2 tumor and non-transgenic mammary control cells. Cells were stimulated with 3.6 μM E2 or TAM and compared to vehicle (EtOH). (C, D) Proliferation at the indicated time points was determined by (C) MTT assays and (D) BrdU incorporation. (E) Colony formation of primary BCL9-2 tumor cells and of non-transgenic control cells cultured on collagen. Shown are the absolute colony numbers of untreated versus E2 and TAM treated primary cells after four days in culture. (F) Representative bright-field images of colonies from primary control and tumor cells treated with the indicated conditions, on day 4 and 10 after culture on collagen. The graphs show the mean of at least three independent experiments and of their standard error, relative to vehicle treated controls. * indicates significant differences for P<.05. Scale bars: 100 μm for bright field in (A) and 200 μm in (F).
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Figure 5: Primary breast tumor cells from BCL9-2 transgenic mice respond to estrogen and tamoxifen(A) Bright field image and co-immunofluorescence stains of primary cells established from mammary tumors of BCL9-2 females. Merged pictures of the co-stains with the indicated antibodies are shown. (B) Dose-response analysis of primary control and of primary BCL9-2 tumor cells for increasing concentrations of estrogen (E2) and tamoxifen (TAM), relative to vehicle treated cells. Cells were stimulated with the indicated concentrations for 48 hours and cell viability was determined by MTT assays. (C-F) Proliferation and colony formation of primary BCL9-2 tumor and non-transgenic mammary control cells. Cells were stimulated with 3.6 μM E2 or TAM and compared to vehicle (EtOH). (C, D) Proliferation at the indicated time points was determined by (C) MTT assays and (D) BrdU incorporation. (E) Colony formation of primary BCL9-2 tumor cells and of non-transgenic control cells cultured on collagen. Shown are the absolute colony numbers of untreated versus E2 and TAM treated primary cells after four days in culture. (F) Representative bright-field images of colonies from primary control and tumor cells treated with the indicated conditions, on day 4 and 10 after culture on collagen. The graphs show the mean of at least three independent experiments and of their standard error, relative to vehicle treated controls. * indicates significant differences for P<.05. Scale bars: 100 μm for bright field in (A) and 200 μm in (F).

Mentions: To assess the growth potential and hormone sensitivity of the BCL9-2 tumors we established primary cell cultures of BCL9-2 breast tumors and of mammary glands from age matched, non-transgenic controls (Fig. 5 and Suppl. Fig. 3). We confirmed by immunofluorescence stains that the primary tumor cells retained all linage specific markers as detected by immunostains on tumor tissue sections (Fig. 5A and Suppl. Fig. 3A). The culture also contained single SMA positive cells. Importantly, primary tumor cells were highly positive for BCL9-2 and ER, which co-localized in the nuclei of primary tumor cells (Fig. 5A).


The BCL9-2 proto-oncogene governs estrogen receptor alpha expression in breast tumorigenesis.

Zatula N, Wiese M, Bunzendahl J, Birchmeier W, Perske C, Bleckmann A, Brembeck FH - Oncotarget (2014)

Primary breast tumor cells from BCL9-2 transgenic mice respond to estrogen and tamoxifen(A) Bright field image and co-immunofluorescence stains of primary cells established from mammary tumors of BCL9-2 females. Merged pictures of the co-stains with the indicated antibodies are shown. (B) Dose-response analysis of primary control and of primary BCL9-2 tumor cells for increasing concentrations of estrogen (E2) and tamoxifen (TAM), relative to vehicle treated cells. Cells were stimulated with the indicated concentrations for 48 hours and cell viability was determined by MTT assays. (C-F) Proliferation and colony formation of primary BCL9-2 tumor and non-transgenic mammary control cells. Cells were stimulated with 3.6 μM E2 or TAM and compared to vehicle (EtOH). (C, D) Proliferation at the indicated time points was determined by (C) MTT assays and (D) BrdU incorporation. (E) Colony formation of primary BCL9-2 tumor cells and of non-transgenic control cells cultured on collagen. Shown are the absolute colony numbers of untreated versus E2 and TAM treated primary cells after four days in culture. (F) Representative bright-field images of colonies from primary control and tumor cells treated with the indicated conditions, on day 4 and 10 after culture on collagen. The graphs show the mean of at least three independent experiments and of their standard error, relative to vehicle treated controls. * indicates significant differences for P<.05. Scale bars: 100 μm for bright field in (A) and 200 μm in (F).
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Figure 5: Primary breast tumor cells from BCL9-2 transgenic mice respond to estrogen and tamoxifen(A) Bright field image and co-immunofluorescence stains of primary cells established from mammary tumors of BCL9-2 females. Merged pictures of the co-stains with the indicated antibodies are shown. (B) Dose-response analysis of primary control and of primary BCL9-2 tumor cells for increasing concentrations of estrogen (E2) and tamoxifen (TAM), relative to vehicle treated cells. Cells were stimulated with the indicated concentrations for 48 hours and cell viability was determined by MTT assays. (C-F) Proliferation and colony formation of primary BCL9-2 tumor and non-transgenic mammary control cells. Cells were stimulated with 3.6 μM E2 or TAM and compared to vehicle (EtOH). (C, D) Proliferation at the indicated time points was determined by (C) MTT assays and (D) BrdU incorporation. (E) Colony formation of primary BCL9-2 tumor cells and of non-transgenic control cells cultured on collagen. Shown are the absolute colony numbers of untreated versus E2 and TAM treated primary cells after four days in culture. (F) Representative bright-field images of colonies from primary control and tumor cells treated with the indicated conditions, on day 4 and 10 after culture on collagen. The graphs show the mean of at least three independent experiments and of their standard error, relative to vehicle treated controls. * indicates significant differences for P<.05. Scale bars: 100 μm for bright field in (A) and 200 μm in (F).
Mentions: To assess the growth potential and hormone sensitivity of the BCL9-2 tumors we established primary cell cultures of BCL9-2 breast tumors and of mammary glands from age matched, non-transgenic controls (Fig. 5 and Suppl. Fig. 3). We confirmed by immunofluorescence stains that the primary tumor cells retained all linage specific markers as detected by immunostains on tumor tissue sections (Fig. 5A and Suppl. Fig. 3A). The culture also contained single SMA positive cells. Importantly, primary tumor cells were highly positive for BCL9-2 and ER, which co-localized in the nuclei of primary tumor cells (Fig. 5A).

Bottom Line: BCL9-2 overexpression in vivo delayed the mammary involution and induced alveolar hyperplasia.We found, that primary cell cultures of BCL9-2 breast tumors responded to tamoxifen treatment.Finally, we describe a novel mechanism, how BCL9-2 regulates ER transcription by interaction with Sp1 through the proximal ESR1 gene promoter.

View Article: PubMed Central - PubMed

Affiliation: Tumor Biology and Signal Transduction, Georg-August-University Göttingen, Germany. Dept. of Hematology and Medical Oncology, Georg-August-University Göttingen, Germany.

ABSTRACT
The majority of human breast cancers express estrogen receptor alpha (ER), which is important for therapy with anti-estrogens. Here we describe the role of BCL9-2, a proto-oncogene previously characterized as co-activator of Wnt/ß-catenin signaling, for mammary tumorigenesis in mice and human. ER positive human breast cancers showed overexpression of BCL9-2 and tamoxifen treated patients with high BCL9-2 demonstrated a better survival. BCL9-2 was upregulated during puberty and pregnancy in normal mammary epithelia, but downregulated in the involuted gland. BCL9-2 overexpression in vivo delayed the mammary involution and induced alveolar hyperplasia. Moreover, aged BCL9-2 transgenic mice developed ductal-like mammary tumors with high nuclear ER expression. We found, that primary cell cultures of BCL9-2 breast tumors responded to tamoxifen treatment. Moreover, BCL9-2 regulated the expression of ER and the proliferation of human breast cancer cells independently of ß-catenin. Finally, we describe a novel mechanism, how BCL9-2 regulates ER transcription by interaction with Sp1 through the proximal ESR1 gene promoter. In summary, BCL9-2 induces ER positive breast cancers in vivo, regulates ER expression by a novel ß-catenin independent mechanism in breast cancer cells, and might predict the therapy response to tamoxifen treatment.

Show MeSH
Related in: MedlinePlus