Transcriptional regulation of human eosinophil RNases by an evolutionary- conserved sequence motif in primate genome

• Published in: BMC molecular biology 2007-10, Vol.8 (1), p.89-89
• Main Authors: Wang, Hsiu-Yu, Chang, Hao-Teng, Pai, Tun-Wen, Wu, Chung-I, Lee, Yuan-Hung, Chang, Yen-Hsin, Tai, Hsiu-Ling, Tang, Chuan-Yi, Chou, Wei-Yao, Chang, Margaret Dah-Tsyr
• Format: Article
• Language: English
• Subjects: Animals; Base Sequence; Binding Sites; Cells, Cultured; Conserved Sequence - physiology; DNA binding proteins; DNA-Binding Proteins - metabolism; Eosinophil Cationic Protein - genetics; Eosinophil-Derived Neurotoxin - genetics; Eosinophils; Eosinophils - enzymology; Evolution, Molecular; Gene Expression Regulation, Enzymologic; Genetic aspects; Genome; Health aspects; Humans; Macaca; Molecular Sequence Data; Phylogeny; Primates; Primates - genetics; Promoter Regions, Genetic; Ribonuclease; Ribonucleases - genetics; Sequence Deletion; Sequence Homology, Nucleic Acid; Sp1 Transcription Factor - metabolism; Transcription Factors - metabolism; Transcription, Genetic
• ISSN: 1471-2199; 1471-2199
• DOI: 10.1186/1471-2199-8-89

Human eosinophil-derived neurotoxin (edn) and eosinophil cationic protein (ecp) are members of a subfamily of primate ribonuclease (rnase) genes. Although they are generated by gene duplication event, distinct edn and ecp expression profile in various tissues have been reported. In this study, we obtained the upstream promoter sequences of several representative primate eosinophil rnases. Bioinformatic analysis revealed the presence of a shared 34-nucleotide (nt) sequence stretch located at -81 to -48 in all edn promoters and macaque ecp promoter. Such a unique sequence motif constituted a region essential for transactivation of human edn in hepatocellular carcinoma cells. Gel electrophoretic mobility shift assay, transient transfection and scanning mutagenesis experiments allowed us to identify binding sites for two transcription factors, Myc-associated zinc finger protein (MAZ) and SV-40 protein-1 (Sp1), within the 34-nt segment. Subsequent in vitro and in vivo binding assays demonstrated a direct molecular interaction between this 34-nt region and MAZ and Sp1. Interestingly, overexpression of MAZ and Sp1 respectively repressed and enhanced edn promoter activity. The regulatory transactivation motif was mapped to the evolutionarily conserved -74/-65 region of the edn promoter, which was guanidine-rich and critical for recognition by both transcription factors. Our results provide the first direct evidence that MAZ and Sp1 play important roles on the transcriptional activation of the human edn promoter through specific binding to a 34-nt segment present in representative primate eosinophil rnase promoters.