Sequences in the A – intergenic region are not required for stage-specific regulation of the human -globin gene locus

The human -globin locus has been extensively studied as a model of tissue and developmental stage-specific gene expression. Structural mapping of naturally occurring mutations, including transfection and transgenic studies, and the recent finding of intergenic transcripts have suggested that there are cis-acting sequence elements in the A – intergenic region involved in regulating and -globin gene expression. To determine whether previously identified sequences in the A – intergenic region are required for appropriate developmental expression of the human -globin gene cluster, transgenic mice were generated by transfer of yeast artificial chromosomes containing the entire human -globin locus. Three different deletions of the A – intergenic region were introduced, including (i) deletion of the 750-bp A 3 regulatory element (A e), (ii) deletion of 3.2 kb upstream of the -globin gene encompassing pyrimidine-rich sequences and the recently described intergenic transcript initiation site, and (iii) deletion of a 12.5-kb fragment encompassing most of the A – globin intergenic region. Analysis of multiple transgenic lines carrying these deletion constructs demonstrated that the normal stage-specific sequential expression of the -, -, and -globin genes was preserved, despite deletion of these putative regulatory sequences. These studies suggest that regulatory sequences required for activation and silencing of the human -globin gene family during ontogeny reside proximally to the genes and immediately 5 to the human and -globin genes.