title: CCAAT/enhancer-binding protein beta inhibits proliferation in monocytic cells by affecting the retinoblastoma protein/E2F/cyclin E pathway but is not directly required for macrophage morphology.
abstract: Monocytic differentiation is orchestrated by complex networks that are not fully understood. This study further elucidates the involvement of transcription factor CCAAT/enhancer-binding protein β (C/EBPβ). Initially, we demonstrated a marked increase in nuclear C/EBPβ-liver-enriched activating protein* (LAP*)/liver-enriched activating protein (LAP) levels and LAP/liver-enriched inhibiting protein (LIP) ratios in phorbol 12-myristate 13-acetate (PMA)-treated differentiating THP-1 premonocytic cells accompanied by reduced proliferation. To directly study C/EBPβ effects on monocytic cells, we generated novel THP-1-derived (low endogenous C/EBPβ) cell lines stably overexpressing C/EBPβ isoforms. Most importantly, cells predominantly overexpressing LAP* (C/EBPβ-long), but not those overexpressing LIP (C/EBPβ-short), exhibited a reduced proliferation, with no effect on morphology. PMA-induced inhibition of proliferation was attenuated in C/EBPβ-short cells. In C/EBPβ(WT) macrophage-like cells (high endogenous C/EBPβ), we measured a reduced proliferation/cycling index compared with C/EBPβ(KO). The typical macrophage morphology was only observed in C/EBPβ(WT), whereas C/EBPβ(KO) stayed round. C/EBPα did not compensate for C/EBPβ effects on proliferation/morphology. Serum reduction, an independent approach known to inhibit proliferation, induced macrophage morphology in C/EBPβ(KO) macrophage-like cells but not THP-1. In PMA-treated THP-1 and C/EBPβ-long cells, a reduced phosphorylation of cell cycle repressor retinoblastoma was found. In addition, C/EBPβ-long cells showed reduced c-Myc expression accompanied by increased CDK inhibitor p27 and reduced cyclin D1 levels. Finally, C/EBPβ-long and C/EBPβ(WT) cells exhibited low E2F1 and cyclin E levels, and C/EBPβ overexpression was found to inhibit cyclin E1 promoter-dependent transcription. Our results suggest that C/EBPβ reduces monocytic proliferation by affecting the retinoblastoma/E2F/cyclin E pathway and that it may contribute to, but is not directly required for, macrophage morphology. Inhibition of proliferation by C/EBPβ may be important for coordinated monocytic differentiation.
authors.affiliation: Institute of Clinical Chemistry, Hannover Medical School, Carl-Neuberg-Strasse 1, 30625 Hannover, Germany.
authors.forename: Romina
authors.initials: R
authors.lastname: Gutsch
authors.forename: Judith D
authors.initials: JD
authors.lastname: Kandemir
authors.forename: Daniel
authors.initials: D
authors.lastname: Pietsch
authors.forename: Christian
authors.initials: C
authors.lastname: Cappello
authors.forename: Johann
authors.initials: J
authors.lastname: Meyer
authors.forename: Kathrin
authors.initials: K
authors.lastname: Simanowski
authors.forename: René
authors.initials: R
authors.lastname: Huber
authors.forename: Korbinian
authors.initials: K
authors.lastname: Brand
mesh_terms: D000818:Animals; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000235:genetics / Q000378:metabolism*; D022762:CCAAT-Enhancer-Binding Proteins / Q000235:genetics / Q000378:metabolism; D002273:Carcinogens / Q000494:pharmacology; D002460:Cell Line; D049109:Cell Proliferation; D019927:Cyclin E / Q000235:genetics / Q000378:metabolism*; D050687:E2F1 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects / Q000235:genetics; D055785:Gene Knockdown Techniques; D006801:Humans; D008264:Macrophages; D051379:Mice; D009000:Monocytes / Q000166:cytology / Q000378:metabolism*; D015513:Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D016160:Retinoblastoma Protein / Q000235:genetics / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
chemical_list: D022782:CCAAT-Enhancer-Binding Protein-beta; D022762:CCAAT-Enhancer-Binding Proteins; C475820:CCNE1 protein, human; C515848:CEBPA protein, human; C530718:CEBPA protein, mouse; C545012:CEBPB protein, human; D002273:Carcinogens; C545013:Cebpb protein, mouse; D019927:Cyclin E; D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; C493754:E2f1 protein, mouse; D015513:Oncogene Proteins; D020033:Protein Isoforms; D016160:Retinoblastoma Protein; D013755:Tetradecanoylphorbol Acetate