Hypoxia Down-regulates CCAAT/Enhancer Binding Protein-α Expression in Breast Cancer Cells

• Published in: Cancer research (Chicago, Ill.) Ill.), 2008-04, Vol.68 (7), p.2158-2165
• Main Authors: SEIFEDDINE, Ramzi, DREIEIN, Anne, BAROUKI, Robert, MASSAAD-MASSADE, Liliane, BLANC, Etienne, FULCHIGNONI-LATAUD, Marie-Claude, LE FRERE BELDA, Marie-Aude, LECURU, Fabrice, HERVEE MAYI, Thérèse, MAZURE, Nathalie, FAVAUDON, Vincent, MASSAAD, Charbel
• Format: Article
• Language: English
• Subjects: Antineoplastic agents; Base Sequence; Biochemistry, Molecular Biology; Biological and medical sciences; Blotting, Western; Breast Neoplasms; Breast Neoplasms - genetics; Breast Neoplasms - metabolism; Breast Neoplasms - pathology; CCAAT-Enhancer-Binding Protein-alpha; CCAAT-Enhancer-Binding Protein-alpha - biosynthesis; CCAAT-Enhancer-Binding Protein-alpha - genetics; CCAAT-Enhancer-Binding Protein-alpha - metabolism; Cell Cycle; Cell Cycle - physiology; Cell Growth Processes; Cell Growth Processes - physiology; Cell Hypoxia; Cell Hypoxia - genetics; Cell Line, Tumor; Deferoxamine; Deferoxamine - pharmacology; Down-Regulation; Electrophoretic Mobility Shift Assay; Gynecology. Andrology. Obstetrics; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Hypoxia-Inducible Factor 1, alpha Subunit - genetics; Hypoxia-Inducible Factor 1, alpha Subunit - metabolism; Immunohistochemistry; Life Sciences; Mammary gland diseases; Medical sciences; Molecular Sequence Data; Pharmacology. Drug treatments; Promoter Regions (Genetics); Promoter Regions, Genetic - drug effects; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; RNA, Messenger - biosynthesis; RNA, Messenger - genetics; RNA, Small Interfering; RNA, Small Interfering - genetics; Subcellular Fractions; Subcellular Fractions - metabolism; Transcription, Genetic; Transcription, Genetic - drug effects; Transfection; Tumors
• ISSN: 0008-5472; 1538-7445
• DOI: 10.1158/0008-5472.CAN-07-1190

The transcription factor CCAAT/enhancer binding protein-alpha (C/EBP alpha) is involved in the control of cell differentiation and proliferation, and has been suggested to act as a tumor suppressor in several cancers. By using microarray analysis, we have previously shown that hypoxia and estrogen down-regulate C/EBP alpha mRNA in T-47D breast cancer cells. Here, we have examined the mechanism by which the down-regulation by hypoxia takes place. Using the specific RNA polymerase II inhibitor 5,6-dichlorobenzimidazole-1-beta-D-ribofuranoside, the mRNA stability was analyzed under normoxia or hypoxia by quantitative reverse transcription-PCR. Hypoxia reduced the half-life of C/EBP alpha mRNA by approximately 30%. C/EBP alpha gene promoter studies indicated that hypoxia also repressed the transcription of the gene and identified a hypoxia-responsive element (-522; -527 bp), which binds to hypoxia-inducible factor (HIF)-1 alpha, as essential for down-regulation of C/EBP alpha transcription in hypoxia. Immunocytochemical analysis showed that C/EBP alpha was localized in the nucleus at 21% O(2), but was mostly cytoplasmic under 1% O(2). Knockdown of HIF-1 alpha by RNAi restored C/EBP alpha to normal levels under hypoxic conditions. Immunohistochemical studies of 10 tumor samples did not show any colocalization of C/EBP alpha and glucose transporter 1 (used as a marker for hypoxia). Taken together, these results show that hypoxia down-regulates C/EBP alpha expression in breast cancer cells by several mechanisms, including transcriptional and posttranscriptional effects. The down-regulation of C/EBP alpha in hypoxia is mediated by HIF-1.