Differential large-scale chromatin compaction and intranuclear positioning of transcribed versus non-transcribed transgene arrays containing beta-globin regulatory sequences.
Previous work has demonstrated a more decondensed large-scale chromatin structure and a more internal nuclear position for gene-rich versus gene-poor chromosome regions. Here, we show that large-scale chromatin opening and changes in intranuclear positioning of chromosome regions can be induced by normal levels of endogenous transcription factors acting on mammalian regulatory sequences. We transfected mouse erythroleukemia cells with a 15 kbp plasmid containing a lac operator repeat plus beta-globin regulatory sequences driving a beta-galactosidase reporter gene. After green-fluorescent-protein/lac-repressor fusion-protein binding or after fluorescence in situ hybridization, the volume and location of the transgene array signal were measured. With both detection methods, we found that the volume was severalfold larger when transcription was on. While silent transgene arrays were located close to the nuclear membrane, we observed a significantly more internal position for the transcriptionally active state. Our results indicate that both large-scale chromatin decondensation and changes in nuclear positioning as observed for large, complex gene-rich chromosome regions can be reproduced by endogenous regulatory sequences acting within simple repetitive transgene arrays.
Department Biologie II, Ludwig-Maximilians-Universität München, Grosshaderner Str. 2, 82152 Martinsried, Germany. dietzel@lmu.de
Steffen
S
Dietzel
Kourosh
K
Zolghadr
Claudia
C
Hepperger
Andrew S
AS
Belmont
D000818:Animals; D002843:Chromatin / Q000235:genetics* / Q000378:metabolism; D042002:Chromatin Assembly and Disassembly / Q000235:genetics / Q000502:physiology; D005786:Gene Expression Regulation / Q000235:genetics* / Q000502:physiology; D017930:Genes, Reporter / Q000235:genetics / Q000502:physiology; D005914:Globins / Q000235:genetics* / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D017404:In Situ Hybridization, Fluorescence; D051379:Mice; D011401:Promoter Regions, Genetic; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics* / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014407:Tumor Cells, Cultured
D002843:Chromatin; D014157:Transcription Factors; D049452:Green Fluorescent Proteins; D005914:Globins