Transcriptional silencing of human Alu sequences and inhibition of protein binding in the box B regulatory elements by 5′‐CG‐3′ methylation

Abstract

In earlier work, we demonstrated that 5'-CG-3' methylation inhibits the transcriptional activity of human Alu elements associated with the alpha 1-globin and the angiogenin genes in a cell-free transcription system from HeLa nuclear extracts. These studies have been extended to different Alu sequences and to investigations on the mechanism involved in transcriptional silencing by methylation. By comparing the results of DNase I and dimethyl sulfate (DMS) in vitro footprinting on a consensus sequence in the RNA polymerase III promoter control B region between the unmethylated and the 5'-CG-3' methylated B box, evidence has been adduced for effects of 5'-CG-3' methylation on the interaction of specific nuclear proteins with DNA sequences in the B control region of the Alu elements. These results are consistent with the interpretation that the 5'-CG-3' methylation interferes with the binding of proteins that are essential for the function of the B control region in these RNA polymerase III-transcribed elements, and that promoter methylation thus inhibits transcription.