Hypoxia down-regulates CCAAT/enhancer binding protein-alpha expression in breast cancer cells.

Abstract

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.

• Authors: Ramzi Seifeddine, Anne Dreiem, Etienne Blanc, Marie-Claude Fulchignoni-Lataud, Marie-Aude Le Frère Belda, Fabrice Lecuru, Thérèse Hervèe Mayi, Nathalie Mazure, Vincent Favaudon, Charbel Massaad, Robert Barouki, Liliane Massaad-Massade
• Journal: Cancer research (Cancer Res) Vol. 68 Issue 7 Pg. 2158-65 (Apr 01 2008) ISSN: 1538-7445 [Electronic] United States
• PMID: 18381421 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• CCAAT-Enhancer-Binding Protein-alpha
• HIF1A protein, human
• Hypoxia-Inducible Factor 1, alpha Subunit
• RNA, Messenger
• RNA, Small Interfering
• Deferoxamine

Topics

• Base Sequence
• Blotting, Western
• Breast Neoplasms (genetics, metabolism, pathology)
• CCAAT-Enhancer-Binding Protein-alpha (biosynthesis, genetics, metabolism)
• Cell Cycle (physiology)
• Cell Growth Processes (physiology)
• Cell Hypoxia (genetics)
• Cell Line, Tumor
• Deferoxamine (pharmacology)
• Down-Regulation
• Electrophoretic Mobility Shift Assay
• Humans
• Hypoxia-Inducible Factor 1, alpha Subunit (genetics, metabolism)
• Immunohistochemistry
• Molecular Sequence Data
• Promoter Regions, Genetic (drug effects)
• RNA, Messenger (biosynthesis, genetics)
• RNA, Small Interfering (genetics)
• Reverse Transcriptase Polymerase Chain Reaction
• Subcellular Fractions (metabolism)
• Transcription, Genetic (drug effects)
• Transfection