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Nucleic Acids Res. 2000 May 1;28(9):1921-8.

MeCP2 driven transcriptional repression in vitro: selectivity for methylated DNA, action at a distance and contacts with the basal transcription machinery.

Author information

1
Laboratory of Molecular Embryology, National Institute of Child Heath and Human Development, NIH, Building 18T, Room 106, Bethesda, MD 20892-5431, USA.

Abstract

The pathways for selective transcriptional repression of methylated DNA templates by the methyl-CpG-binding protein MeCP2 have been investigated using a purified in vitro transcription system that does not assemble chromatin. MeCP2 selectively inhibits transcription complex assembly on methylated DNA but does not destabilize a pre-assembled transcription complex. MeCP2 functions to repress transcription at a distance of >500 bp from the transcription start site. The transcription repression domain (TRD) of MeCP2 will repress transcription in vitro when fused to a heterologous Gal4 DNA-binding domain. The TRD associates with TFIIB. Exogenous TFIIB does not relieve transcriptional repression established by either intact MeCP2 or a Gal4-TRD fusion protein under these in vitro conditions, nor does the addition of histone deacetylase inhibitors. We find that the transcriptional repression established by both MeCP2 and the Gal4-TRD fusion protein in vitro also correlates with selective assembly of large nucleoprotein complexes. The formation of such complexes reflects a local concentration of DNA-bound transcriptional repressor that may stabilize a state of repression even in the presence of exogenous transcriptional machinery.

PMID:
10756192
PMCID:
PMC103274
DOI:
10.1093/nar/28.9.1921
[Indexed for MEDLINE]
Free PMC Article

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