Nuclear protein factors and erythroid transcription of the human A gamma-globin gene

We have used DNaseI footprinting and gel mobility assays to analyze the upstream region of the human A gamma-globin gene promoter. Four protein factors were found to bind this region. A non-erythroid factor present in the 0.4M KCl fraction of a heparin agarose column binds to the CAC box (-140). A ubiquitous octamer factor present in the 0.2M fraction binds to an ATGCAAT element (-175), but is completed out by the erythroid specific factor NF-E1 (in the 0.4M KCl fraction), which binds a site (-186) immediately flanking the octamer. A novel factor binding to a stretch of 8A around -233, was identified in the 0.2M KCl fraction. This factor is not present in HeLa nuclear extracts. To study the transcriptional importance of these protein binding sites we have used an "A gamma-minilocus", similar to that described for the beta-globin gene (1) in K562 cells. This provides evidence that the NF-E1 and CAC box in the -210 to -122 region of the A gamma-promoter are important for the efficient expression of the gamma-globin gene.