Highly compacted chromatin formed in vitro reflects the dynamics of transcription activation in vivo.
High-order chromatin was reconstituted in vitro. This species reflects the criteria associated with transcriptional regulation in vivo. Histone H1 was determinant to formation of condensed structures, with deacetylated histones giving rise to highly compacted chromatin that approximated 30 nm fibers as evidenced by electron microscopy. Using the PEPCK promoter, we validated the integrity of these templates that were refractory to transcription by attaining transcription through the progressive action of the pertinent factors. The retinoic acid receptor binds to highly compacted chromatin, but the NF1 transcription factor binds only after histone acetylation by p300 and SWI/SNF-mediated nucleosome mobilization, reflecting the in vivo case. Mapping studies revealed the same pattern of nucleosomal repositioning on the PEPCK promoter in vitro and in vivo, correlating with NF1 binding and transcription. The reconstitution of such highly compacted "30 nm" chromatin that mimics in vivo characteristics should advance studies of its conversion to a transcriptionally active form.
Li
Guohong
G
Department of Biochemistry, New York University School of Medicine, 522 First Avenue, New York, NY 10016, USA.
Margueron
Raphael
R
Hu
Guobin
G
Stokes
David
D
Wang
Yuh-Hwa
YH
Reinberg
Danny
D
D000107:Acetylation; D002843:Chromatin / Q000378:metabolism / Q000648:ultrastructure; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006657:Histones / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D051839:NFI Transcription Factors / Q000235:genetics / Q000378:metabolism; D009707:Nucleosomes / Q000378:metabolism; D010729:Phosphoenolpyruvate Carboxykinase (GTP) / Q000235:genetics; D011401:Promoter Regions, Genetic; D018168:Receptors, Retinoic Acid / Q000235:genetics / Q000378:metabolism; D047488:Retinoid X Receptors / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D015533:Transcriptional Activation; D050880:p300-CBP Transcription Factors / Q000235:genetics / Q000378:metabolism; D000099224:p300-CBP-Associated Factor
D002843:Chromatin; D002868:Chromosomal Proteins, Non-Histone; D006657:Histones; D051839:NFI Transcription Factors; D009707:Nucleosomes; D010729:Phosphoenolpyruvate Carboxykinase (GTP); D018168:Receptors, Retinoic Acid; D047488:Retinoid X Receptors; D014157:Transcription Factors; D050880:p300-CBP Transcription Factors; D000099224:p300-CBP-Associated Factor; C445179:SWI-SNF-B chromatin-remodeling complex
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