The CCAAT/Enhancer-Binding Protein Beta-2 Isoform (CEBPβ-2) Upregulates Galectin-7 Expression in Human Breast Cancer Cells

Galectin-7 is considered a gene under the control of p53. However, elevated expression of galectin-7 has been reported in several forms of cancer harboring an inactive p53 pathway. This is especially true for breast cancer where galectin-7 expression is readily expressed in a high proportion in basal-like breast cancer tissues, conferring cancer cells with increased resistance to cell death and metastatic properties. These observations suggest that other transcription factors are capable of inducing galectin-7 expression. In the present work, we have examined the role of CCAAT/enhancer-binding protein beta (C/EBPβ) in inducing expression of galectin-7. C/EBP proteins have been shown to contribute to breast cancer by upregulating pro-metastatic genes. We paid particular attention to C/EBPβ-2 (also known as LAP2), the most transcriptionally active of the C/EBPβ isoforms. Our results showed that ectopic expression of C/EBPβ-2 in human breast cancer cells was sufficient to induce expression of galectin-7 at both the mRNA and protein levels. In silico analysis further revealed the presence of an established CEBP element in the galectin-7 promoter. Mutation of this binding site abolished the transcriptional activity of the galectin-7 promoter. Chromatin immunoprecipitation analysis confirmed that C/EBPβ-2 binds to the endogenous galectin-7 promoter. Analysis of galectin-7 protein expression in normal epithelia and in breast carcinoma by immunohistochemistry further showed the expression pattern of C/EBPβ closely micmicked that of galectin-7, most notably in mammary myoepithelial cells and basal-like breast cancer where galectin-7 is preferentially expressed. Taken together, our findings suggest that C/EBPβ is an important mediator of galectin-7 gene activation in breast cancer cells and highlight the different transcriptional mechanisms controlling galectin-7 in cancer cells.

[1]  L. Haoran,et al.  Transcriptome Analysis of the Trachinotus ovatus: Identification of Reproduction, Growth and Immune-Related Genes and Microsatellite Markers , 2014, PloS one.

[2]  Alan Mackay,et al.  Expression profiling of purified normal human luminal and myoepithelial breast cells: identification of novel prognostic markers for breast cancer. , 2004, Cancer research.

[3]  K. Kinzler,et al.  A model for p53-induced apoptosis , 1997, Nature.

[4]  R. Smart,et al.  CCAAT/enhancer binding protein-β is a mediator of keratinocyte survival and skin tumorigenesis involving oncogenic Ras signaling , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[5]  T. Magnaldo,et al.  Galectin-7, a human 14-kDa S-lectin, specifically expressed in keratinocytes and sensitive to retinoic acid. , 1995, Developmental biology.

[6]  C. Zahnow,et al.  Overexpression of C/EBPbeta-LIP, a naturally occurring, dominant-negative transcription factor, in human breast cancer. , 1997, Journal of the National Cancer Institute.

[7]  Giovanni Parmigiani,et al.  A primary xenograft model of small-cell lung cancer reveals irreversible changes in gene expression imposed by culture in vitro. , 2009, Cancer research.

[8]  Andy J. Minn,et al.  Genes that mediate breast cancer metastasis to lung , 2005, Nature.

[9]  E. Eaton,et al.  Characterization of C/EBPβ isoforms in normal versus neoplastic mammary epithelial cells , 2001 .

[10]  B. Dörken,et al.  C/EBPbeta regulates transcription factors critical for proliferation and survival of multiple myeloma cells. , 2009, Blood.

[11]  C. Zahnow,et al.  Overexpression of C/EBPβ-LIP, a Naturally Occurring, Dominant-Negative Transcription Factor, in Human Breast Cancer , 1997 .

[12]  Kay Nieselt,et al.  Progression-specific genes identified by expression profiling of matched ductal carcinomas in situ and invasive breast tumors, combining laser capture microdissection and oligonucleotide microarray analysis. , 2006, Cancer research.

[13]  H. Nishiyama,et al.  Sensitizing effect of galectin-7 in urothelial cancer to cisplatin through the accumulation of intracellular reactive oxygen species. , 2007, Cancer research.

[14]  L. Gaboury,et al.  A Mutation in the Carbohydrate Recognition Domain Drives a Phenotypic Switch in the Role of Galectin-7 in Prostate Cancer , 2015, PloS one.

[15]  J. Hébert,et al.  Galectin-7 in lymphoma: elevated expression in human lymphoid malignancies and decreased lymphoma dissemination by antisense strategies in experimental model. , 2007, Cancer research.

[16]  M. Lazar,et al.  Retinoic acid blocks adipogenesis by inhibiting C/EBPbeta-mediated transcription , 1997, Molecular and cellular biology.

[17]  C. Zahnow,et al.  CCAAT/enhancer-binding protein β: its role in breast cancer and associations with receptor tyrosine kinases , 2009, Expert Reviews in Molecular Medicine.

[18]  Qing‐Yu He,et al.  Proteomics of buccal squamous cell carcinoma: The involvement of multiple pathways in tumorigenesis , 2004, Proteomics.

[19]  S. Barondes,et al.  Galectins. Structure and function of a large family of animal lectins. , 1994, The Journal of biological chemistry.

[20]  J. Rosen,et al.  C/EBPβ (CCAAT/Enhancer Binding Protein) Controls Cell Fate Determination during Mammary Gland Development , 2000 .

[21]  K. Milde-Langosch,et al.  Expression of the CCAAT/Enhancer-Binding Proteins C/EBPα, C/EBPβ and C/EBPδ in Breast Cancer: Correlations with Clinicopathologic Parameters and Cell-Cycle Regulatory Proteins , 2003, Breast Cancer Research and Treatment.

[22]  T. Magnaldo,et al.  Galectin-7, a marker of all types of stratified epithelia. , 1998, Differentiation; research in biological diversity.

[23]  A. V. Timoshenko.  Towards molecular mechanisms regulating the expression of galectins in cancer cells under microenvironmental stress conditions , 2015, Cellular and Molecular Life Sciences.

[24]  G. Lavoie,et al.  Expression of Galectin-7 Is Induced in Breast Cancer Cells by Mutant p53 , 2013, PloS one.

[25]  Brian Raught,et al.  C/EBPβ, but not C/EBPα, is essential for ductal morphogenesis, lobuloalveolar proliferation, and functional differentiation in the mouse mammary gland , 1998 .

[26]  B. Raught,et al.  C/EBPbeta, but not C/EBPalpha, is essential for ductal morphogenesis, lobuloalveolar proliferation, and functional differentiation in the mouse mammary gland. , 1998, Genes & development.

[27]  R. Smart,et al.  C/EBPβ Modulates the Early Events of Keratinocyte Differentiation Involving Growth Arrest and Keratin 1 and Keratin 10 Expression , 1999, Molecular and Cellular Biology.

[28]  T. Löning,et al.  Expression of the CCAAT/enhancer-binding proteins C/EBPalpha, C/EBPbeta and C/EBPdelta in breast cancer: correlations with clinicopathologic parameters and cell-cycle regulatory proteins. , 2003, Breast cancer research and treatment.

[29]  C. Zahnow,et al.  A role for CCAAT/enhancer binding protein beta-liver-enriched inhibitory protein in mammary epithelial cell proliferation. , 2001, Cancer research.

[30]  Edgar Wingender,et al.  PC-TraFF: identification of potentially collaborating transcription factors using pointwise mutual information , 2015, BMC Bioinformatics.

[31]  Jonathan M. Lee,et al.  CCAAT/Enhancer binding protein beta abrogates retinoic acid-induced osteoblast differentiation via repression of Runx2 transcription. , 2007, Molecular endocrinology.

[32]  Christian A. Rees,et al.  Molecular portraits of human breast tumours , 2000, Nature.

[33]  D. Ramji,et al.  CCAAT/enhancer-binding proteins: structure, function and regulation. , 2002, The Biochemical journal.

[34]  H. Thompson,et al.  Gene expression changes associated with chemically induced rat mammary carcinogenesis , 1997, Molecular carcinogenesis.

[35]  E. Eaton,et al.  Characterization of C/EBPbeta isoforms in normal versus neoplastic mammary epithelial cells. , 2001, Journal of cellular physiology.

[36]  J. Rosen,et al.  C/EBPbeta (CCAAT/enhancer binding protein) controls cell fate determination during mammary gland development. , 2000, Molecular endocrinology.

[37]  Louis Gaboury,et al.  Tumorigenesis and Neoplastic Progression Overexpression of Galectin-7 , A Myoepithelial Cell Marker , Enhances Spontaneous Metastasis of Breast Cancer Cells , 2010 .

[38]  M. O’Connor-McCourt,et al.  Transforming growth factor-β1 is the predominant isoform required for breast cancer cell outgrowth in bone , 2009, Oncogene.

[39]  R. Kiss,et al.  Changes in Galectin-7 and Cytokeratin-19 Expression during the Progression of Malignancy in Thyroid Tumors: Diagnostic and Biological Implications , 2002, Modern Pathology.

[40]  Xiao-Jun Ma,et al.  Gene expression profiling of the tumor microenvironment during breast cancer progression , 2009, Breast Cancer Research.

[41]  Carsten O. Peterson,et al.  Estrogen receptor status in breast cancer is associated with remarkably distinct gene expression patterns. , 2001, Cancer research.

[42]  L. Bundy,et al.  CCAAT/enhancer binding protein beta (C/EBPβ)-2 transforms normal mammary epithelial cells and induces epithelial to mesenchymal transition in culture , 2003, Oncogene.

[43]  L. Hennighausen,et al.  The C/EBPbeta transcription factor regulates epithelial cell proliferation and differentiation in the mammary gland. , 1998, Genes & development.

[44]  Shridar Ganesan,et al.  X chromosomal abnormalities in basal-like human breast cancer. , 2006, Cancer cell.

[45]  A. Sarasin,et al.  Galectin-7 overexpression is associated with the apoptotic process in UVB-induced sunburn keratinocytes. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[46]  L. Tan,et al.  Identification of galectin-7 as a potential biomarker for esophageal squamous cell carcinoma by proteomic analysis , 2010, BMC Cancer.

[47]  J. Hirabayashi,et al.  Galectins: a family of animal lectins that decipher glycocodes. , 1996, Journal of biochemistry.