Enhancer-Dependent Transcriptional Oscillations in Mouse Erythroleukemia Cells

Abstract

By using recombinase-mediated cassette exchange, a method that allows integration of single copies of different constructs at the same predetermined chromosomal location, several expression cassettes have been integrated at a randomly chosen locus in the genome of mouse erythroleukemia cells. The cassettes studied contain the human β-globin promoter fused to lacZ coding sequences either alone or linked to DNase I-hypersensitive site HS2, HS3, or HS234 (a large locus control region fragment containing HS2, HS3, and HS4) of the human β-globin locus control region. Analysis of expression of these cassettes revealed mosaic expression patterns reminiscent of, but clearly different from, position effect variegation. Further investigations demonstrated that these mosaic expression patterns are caused by dynamic activation and inactivation of the transcription unit, resulting in oscillations of expression. These oscillations occur once in every few cell cycles at a rate specific for the enhancer present at the locus. DNase I sensitivity studies revealed that the chromatin is accessible and that DNase-hypersensitive sites were present whether or not the transcription unit is active, suggesting that the oscillations occur between transcriptionally competent and transcriptionally active chromatin conformations, rather than between open and closed chromatin conformations. Treatment of oscillating cells with trichostatin A eliminates the oscillations only after the cells have passed through late G₁ or early S, suggesting that these oscillations might be caused by changes in histone acetylation patterns.

ACKNOWLEDGMENTS

We are very grateful to Ronald Nagel for ongoing support of this project; to Ilia Rochlin, Antonietta Marmorato, and Maria Martinovski for excellent technical help; and to E. Klein-Bouhassira, A. Eisen, S. Fiering, A. Skoultchi, C. Schildkraut, and M. Rapport for critical review of the manuscript.

This work was supported by grants NIH HL38655 and HL 55435 from the NIH and by grant-in-aid 95015110 from the American Heart Association.