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Published 18 August 2003. doi:10.1083/jcb.200305013
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© The Rockefeller University Press, 0021-9525/2003/8/565 $5.00
The Journal of Cell Biology, Volume 162, Number 4, 565-574

Article

Visualization of chromatin domains created by the gypsy insulator of Drosophila



Keith Byrd and Victor G. Corces

Department of Biology, Johns Hopkins University, Baltimore, MD 21218

Address correspondence to Victor G. Corces, Department of Biology, Johns Hopkins University, 3400 N. Charles St., Baltimore, MD 21218. Tel.: (410) 516-8749. Fax: (410) 516-5456. email: corces{at}jhu.edu

Insulators might regulate gene expression by establishing and maintaining the organization of the chromatin fiber within the nucleus. Biochemical fractionation and in situ high salt extraction of lysed cells show that two known protein components of the gypsy insulator are present in the nuclear matrix. Using FISH with DNA probes located between two endogenous Su(Hw) binding sites, we show that the intervening DNA is arranged in a loop, with the two insulators located at the base. Mutations in insulator proteins, subjecting the cells to a brief heat shock, or destruction of the nuclear matrix lead to disruption of the loop. Insertion of an additional gypsy insulator in the center of the loop results in the formation of paired loops through the attachment of the inserted sequences to the nuclear matrix. These results suggest that the gypsy insulator might establish higher-order domains of chromatin structure and regulate nuclear organization by tethering the DNA to the nuclear matrix and creating chromatin loops.

Key Words: insulator; chromatin; transcription; nucleus; retrotransposon

Abbreviations used in this paper: ct, cut; MAR, matrix attachment region; NPC, nuclear pore complex; SAR, scaffold attachment region; SCS, specialized chromatin structures; Ubx, Ultrabithorax.


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