Identification of functional CCAAT/enhancer-binding protein and Ets protein binding sites in the human chorionic somatomammotropin enhancer sequences

• Published in: Journal of molecular endocrinology Vol. 47; no. 2; pp. 179 - 193
• Main Authors: Lytras, Aristides, Detillieux, Karen, Cattini, Peter A
• Format: Journal Article
• Language: English
• Published: England Society for Endocrinology 01.10.2011; BioScientifica
• Subjects: Binding Sites; CCAAT-Enhancer-Binding Proteins - metabolism; Cell Line, Tumor; Chromatin Immunoprecipitation; Electrophoretic Mobility Shift Assay; Enhancer Elements, Genetic - genetics; Humans; Placental Lactogen - genetics; Promoter Regions, Genetic - genetics; Protein Binding - genetics; Proto-Oncogene Proteins c-ets - metabolism; Regular papers
• ISSN: 0952-5041; 1479-6813
• DOI: 10.1530/JME-11-0025

The human chorionic somatomammotropin (CS) A and B genes (listed as CSH1 and CSH2 in the HUGO database) are highly expressed in placenta. A 241 bp potent enhancer, nucleotides (nts) 1–241, located at the 3′ end of the CS-B gene (CS-Benh) stimulates promoter activity specifically in placental trophoblast cells in vitro. Strong activity is exerted by a 23 bp element within the CS-Benh (nts 117–139), shown to interact with transcription enhancer factor (TEF) members of the transcription enhancer activator (TEA) DNA-binding domain-containing family. An identical TEF element is present in the homologous (97.5%) CS-Aenh; however, a few nucleotide differences suppress its activity. Previously, we identified regulatory sequences distinct from the TEF element within an 80 bp modulatory domain (nts 1–80) in the CS-Benh. Using structural and functional assays we now show that CCAAT/enhancer-binding protein (C/EBP) binding sites exist in the 80 bp modulatory domains of both enhancers, and an Elk-1 binding site exists in the modulatory domain of the CS-Aenh. C/EBPα or C/EBPβ strongly repressed CSp.CAT activity but stimulated CSp.CAT.CS-Benh activity. In contrast, the equivalent CS-A enhancer sequences were unable to relieve promoter repression. Elk-1 overexpression also resulted in differential effects on the CS-Aenh versus CS-Benh. Finally, we provide evidence for the association of C/EBPβ with the CS-A and CS-B genes in human placental chromatin, including differential involvement of C/EBPβ with the CS-Aenh versus the CS-Benh, and therefore consistent with the notion that these are regions of regulatory significance in vivo. We conclude that members of the C/EBP and Ets families can differentially modulate CS-Benh and CS-Aenh activity.