The human beta-globin locus control region (LCR) consists of four 
erythroid-specific DNaseI hypersensitive sites (HSs) at the 5' end of the 
beta-globin cluster. The LCR functions over a long distance on chromosome 11 to 
regulate transcription and replication of the beta-globin genes. To determine 
whether the HSs function independently or as an integrated unit, we analyzed the 
requirements for long-range transcriptional activation. If the HSs function 
independently, individual HSs would be expected to have long-range activity. In 
contrast, if long-range activity requires multiple HSs, individual HSs would 
have a limited functional distance. HS2, HS3, and a miniLCR containing multiple 
HSs, were separated from a gamma-globin promoter by fragments of phage lambda 
DNA. After stable transfection into K562 cells, HS2 had strong enhancer 
activity, but only when positioned close to the promoter. HS3 also had strong 
enhancer activity, although it was weaker than HS2 and more sensitive to the 
spacer DNA. The miniLCR had the strongest enhancer activity and functioned even 
at a distance of 7.3 kb. A model is proposed in which synergistic interactions 
between HSs confer long-range activation by creating a stable LCR nucleoprotein 
structure, which is competent for recruiting chromatin-modifying enzymes. These 
enzymes would mediate the well-characterized activity of the LCR to modulate 
chromatin structure.