Dataset Information

DNA sequence encoded repression of rRNA gene transcription in chromatin.

ABSTRACT: Eukaryotic genomes are packaged into nucleosomes that occlude DNA from interacting with most DNA-binding proteins. Nucleosome positioning and chromatin organization is critical for gene regulation. We have investigated the mechanism by which nucleosomes are positioned at the promoters of active and silent rRNA genes (rDNA). The reconstitution of nucleosomes on rDNA results in sequence-dependent nucleosome positioning at the rDNA promoter that mimics the chromatin structure of silent rRNA genes in vivo, suggesting that active mechanisms are required to reorganize chromatin structure upon gene activation. Nucleosomes are excluded from positions observed at active rRNA genes, resulting in transcriptional repression on chromatin. We suggest that the repressed state is the default chromatin organization of the rDNA and gene activation requires ATP-dependent chromatin remodelling activities that move the promoter-bound nucleosome about 22-bp upstream. We suggest that nucleosome remodelling precedes promoter-dependent transcriptional activation as specific inhibition of ATP-dependent chromatin remodelling suppresses the initiation of RNA Polymerase I transcription in vitro. Once initiated, RNA Polymerase I is capable of elongating through reconstituted chromatin without apparent displacement of the nucleosomes. The results reveal the functional cooperation of DNA sequence and chromatin remodelling complexes in nucleosome positioning and in establishing the epigenetic active or silent state of rRNA genes.

SUBMITTER: Felle M

PROVIDER: S-EPMC2938192 | biostudies-literature | 2010 Sep

REPOSITORIES: biostudies-literature

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DNA sequence encoded repression of rRNA gene transcription in chromatin.

Felle Max M Exler Josef H JH Merkl Rainer R Dachauer Karoline K Brehm Alexander A Grummt Ingrid I Längst Gernot G

Nucleic acids research 20100425 16

Eukaryotic genomes are packaged into nucleosomes that occlude DNA from interacting with most DNA-binding proteins. Nucleosome positioning and chromatin organization is critical for gene regulation. We have investigated the mechanism by which nucleosomes are positioned at the promoters of active and silent rRNA genes (rDNA). The reconstitution of nucleosomes on rDNA results in sequence-dependent nucleosome positioning at the rDNA promoter that mimics the chromatin structure of silent rRNA genes i ...[more]

PMID: 20421213

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Project description:Many DNA hypermethylated and epigenetically silenced genes in adult cancers are Polycomb group (PcG) marked in embryonic stem (ES) cells. We show that a large region upstream ( approximately 30 kb) of and extending approximately 60 kb around one such gene, GATA-4, is organized-in Tera-2 undifferentiated embryonic carcinoma (EC) cells-in a topologically complex multi-loop conformation that is formed by multiple internal long-range contact regions near areas enriched for EZH2, other PcG proteins, and the signature PcG histone mark, H3K27me3. Small interfering RNA (siRNA)-mediated depletion of EZH2 in undifferentiated Tera-2 cells leads to a significant reduction in the frequency of long-range associations at the GATA-4 locus, seemingly dependent on affecting the H3K27me3 enrichments around those chromatin regions, accompanied by a modest increase in GATA-4 transcription. The chromatin loops completely dissolve, accompanied by loss of PcG proteins and H3K27me3 marks, when Tera-2 cells receive differentiation signals which induce a approximately 60-fold increase in GATA-4 expression. In colon cancer cells, however, the frequency of the long-range interactions are increased in a setting where GATA-4 has no basal transcription and the loops encompass multiple, abnormally DNA hypermethylated CpG islands, and the methyl-cytosine binding protein MBD2 is localized to these CpG islands, including ones near the gene promoter. Removing DNA methylation through genetic disruption of DNA methyltransferases (DKO cells) leads to loss of MBD2 occupancy and to a decrease in the frequency of long-range contacts, such that these now more resemble those in undifferentiated Tera-2 cells. Our findings reveal unexpected similarities in higher order chromatin conformation between stem/precursor cells and adult cancers. We also provide novel insight that PcG-occupied and H3K27me3-enriched regions can form chromatin loops and physically interact in cis around a single gene in mammalian cells. The loops associate with a poised, low transcription state in EC cells and, with the addition of DNA methylation, completely repressed transcription in adult cancer cells.

Dataset Information

DNA sequence encoded repression of rRNA gene transcription in chromatin.

Publications

DNA sequence encoded repression of rRNA gene transcription in chromatin.

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