Intragenic DNA methylation alters chromatin structure and elongation efficiency in mammalian cells.
Transcriptional silencing in mammals is often associated with promoter methylation. However, a considerable number of genomic methylated CpGs exist in transposable elements, which are frequently found in intronic regions. To determine whether intragenic methylation influences transcription efficiency, we used the Cre/loxP-based system, RMCE, to introduce a transgene, methylated exclusively in a region downstream of the promoter, into a specific genomic site. This methylation pattern was maintained in vivo, and yielded a clear decrease in transgene expression relative to an unmethylated control. Notably, RNA polymerase II (Pol II) was depleted exclusively in the methylated region, as was histone H3 di- and trimethylated on Lys4 and acetylated on Lys9 and Lys14. As the methylated region adopts a closed chromatin structure in vivo, we propose that dense intragenic DNA methylation in mammalian cells initiates formation of a chromatin structure that reduces the efficiency of Pol II elongation.
Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, Washington 98109, USA. mlorincz@fhcrc.org
Matthew C
MC
Lorincz
David R
DR
Dickerson
Mike
M
Schmitt
Mark
M
Groudine
D000818:Animals; D015139:Blotting, Southern; D002467:Cell Nucleus / Q000378:metabolism; D002843:Chromatin / Q000737:chemistry*; D047369:Chromatin Immunoprecipitation; D019175:DNA Methylation; D005091:Exons; D005434:Flow Cytometry; D020868:Gene Silencing; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006657:Histones / Q000737:chemistry; D006801:Humans; D007438:Introns; D008239:Lysine / Q000737:chemistry; D008957:Models, Genetic; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013447:Sulfites / Q000494:pharmacology; D014158:Transcription, Genetic; D019076:Transgenes
D002843:Chromatin; D006657:Histones; D013447:Sulfites; D049452:Green Fluorescent Proteins; D008239:Lysine; C009279:sodium bisulfite