THE HUMAN ERYTHROPOIETIN-ENCODING GENE CONTAINS A CAAT BOX, TATA BOXES AND OTHER TRANSCRIPTIONAL REGULATORY ELEMENTS IN ITS 5' FLANKING REGION

We have reported the cloning and expression of a human erythropoietin (hEp)-encoding cDNA [Lee-Huang, Proc. Natl. Acad. Sci. USA 81 (1984) 2708-2712]. Using this hEp cDNA as a probe, we isolated a 9.3-kb BamHI genomic Ep clone from a human leukocyte library soon thereafter. The size and restriction map of this clone is in agreement with restriction analysis of human genomic DNA probed with the hEp cDNA, demonstrating that this clone is representative of the single hEp gene. This clone is unique in that it extends beyond any reported hEp genomic clone by 3.9 kb on the 5' side and by 1.8 kb on the 3' side. The promoter function of the newly described 5' flanking region has been demonstrated by the expression of biologically active hEp in transfected cells. We find that, despite reports to the contrary, hEp does contain classic canonical TATA boxes and a CAAT box. The 5'-flanking region also contains cytokine-responsive consensus sequences, tissue-specific and metal-responsive elements, CRE and GRE sites, and binding sites for transcription factors, including AP1, NF-kappabeta and Spl. These regulatory elements have not been found in the hEp genomic clones thus far reported. The identification of these elements and their precise localization in hEp should be useful in studying the regulation of hEp expression, as well as in gene therapy and physiologic modulation of this hormone.