GATA-1 Directly Induces Physical Proximity between the LCR and β Globin Promoter in Erythroid Cells.

The regulation of higher-order chromatin folding is a poorly understood aspect of nuclear biology. The beta globin locus is one of the most thoroughly studied mammalian loci with regard to its chromatin organization and sub-nuclear localization. The LCR (locus control region) is a powerful enhancer that acts at great distances to regulate the expression of all beta-like globin genes. Recent evidence suggests that the LCR is in physical proximity with the active but not inactive globin gene promoters, thereby looping out up to tens of kilobases of intervening sequence. The LCR-promoter interaction is dynamically regulated throughout development and cellular differentiation. However, the transcription factors that mediate long range “loops” are unknown. Here we examined the essential erythroid transcription factor GATA-1 for its role in LCR-promoter communication using the 3C (Chromosome Conformation Capture) technique. For our studies we used a genetic complementation assay in which erythroid maturation and globin gene expression depend on the activity of a conditional, drug-activated form of GATA-1. The results suggest that GATA-1 induces proximity of the LCR with the active beta-globin promoter. Kinetic studies revealed that the LCR-promoter interaction occurs rapidly and coincides with or even precedes the onset of globin gene transcription. Moreover, “loop” formation correlates with GATA-1 occupancy as determined by ChIP assay, and occurs in the presence of cyclohexamide, indicating that the effects of GATA-1 are direct. Together, these data suggest GATA-1 is a required component for the formation of a tissue-specific chromatin loop. Additional experiments with mutant versions of GATA-1 that address which known GATA-1 cofactors are required for this novel function of GATA-1 will be discussed.