losartan-potassium and cytidylyl-3--5--guanosine

The human erythropoietin gene is expressed predominantly in the kidney and liver in response to hypoxia. Although the signaling cascade for hypoxia is present in many different cell types, the expression of erythropoietin is restricted to only a few tissues. The authors show that the promoter and 5'-untranslated region (5'-UTR) of the erythropoietin gene comprise a CpG island and that methylation of the CpG island correlates inversely with expression. Methylation represses the expression of the erythropoietin gene in 2 ways: high-density methylation of the 5'-UTR recruits a methyl-CpG binding protein to the promoter, and methylation of CpGs in the proximal promoter blocks the association of nuclear proteins. (Blood. 2000;95:111-119)

Topics: 5' Untranslated Regions; Base Sequence; Dinucleoside Phosphates; DNA; DNA Methylation; Erythropoietin; Gene Expression Regulation; HeLa Cells; Humans; Luciferases; Nucleic Acid Conformation; Polymerase Chain Reaction; Promoter Regions, Genetic; Recombinant Proteins; Restriction Mapping; Transfection; Tumor Cells, Cultured

The hypoxia-inducible factor-1 (HIF-1) is a transcriptional activator involved in the expression of oxygen-regulated genes such as that for erythropoietin. Following exposure to low oxygen partial pressure (hypoxia), HIF-1 binds to an hypoxia-response element located 3' to the erythropoietin gene and confers activation of erythropoietin expression. The conserved core HIF-1 binding site (HBS) of the erythropoietin 3' enhancer (CGTG) contains a CpG dinucleotide known to be a potential target of cytosine methylation. We found that methylation of the HBS abolishes HIF-1 DNA binding as well as hypoxic reporter gene activation, suggesting that a methylation-free HBS is mandatory for HIF-1 function. The in vivo methylation pattern of the erythropoietin 3' HBS in various human cell lines and mouse organs was assessed by genomic Southern blotting using a methylation-sensitive restriction enzyme. Whereas this site was essentially methylation-free in the erythropoietin-producing cell line Hep3B, a direct correlation between erythropoietin protein expression and the degree of erythropoietin 3' HBS methylation was found in different HepG2 sublines. However, the finding that this site is partially methylation-free in human cell lines and mouse tissues that do not express erythropoietin suggests that there might be a general selective pressure to keep this site methylation-free, independent of erythropoietin expression.

Topics: Animals; Binding Sites; Carcinoma, Hepatocellular; Cell Hypoxia; Cell Nucleus; Dinucleoside Phosphates; DNA Methylation; DNA-Binding Proteins; Erythropoietin; Gene Expression Regulation; Gene Expression Regulation, Neoplastic; Genes, Reporter; HeLa Cells; Humans; Hypoxia-Inducible Factor 1; Hypoxia-Inducible Factor 1, alpha Subunit; Kidney; L Cells; Leukemia; Liver; Liver Neoplasms; Luciferases; Mice; Neuroblastoma; Nuclear Proteins; Organ Specificity; Recombinant Fusion Proteins; Transcription Factors; Transcriptional Activation; Transfection; Tumor Cells, Cultured