Using conventional and scanning transmission electron microscopy, we have examined the physical basis of long-range enhancer effects between distal and proximal elements in a eukaryotic promoter. Specifically, we have studied binding of human transcription factor Sp1 to 10-base-pair G+C-rich elements ("GC boxes") located at -100 and +1700 relative to the RNA start site. It was previously observed that the distantly located site functions in synergism with the promoter-proximal site to strongly activate transcription in vivo. Here we demonstrate that this synergism is likely to be a direct consequence of interactions between remote and local Sp1, the remote Sp1 translocated to the promoter by a DNA loop. Scanning transmission electron microscopy shows that Sp1 initially forms a tetramer and subsequently assembles multiple tetramers stacked in register at the DNA loop juncture. This unexpected finding not only provides the physical basis for loop formation but also defines a biological process leading to strongly increased concentration of activator protein at the promoter. The mechanism may unify the problem of transcriptional activation by removing enhancer action as a separate class of regulatory activity.

Mastrangelo, I. A., Courey, A. J., Wall, J. S., Jackson, S. P., & Hough, P. V. (1991). DNA looping and Sp1 multimer links: a mechanism for transcriptional synergism and enhancement. Proceedings of the National Academy of Sciences, 88(13), 5670–5674.