Induction of DNA methylation and gene silencing by short interfering RNAs in human cells.
Double-stranded RNAs (dsRNAs) induce post-transcriptional gene silencing in several species of animal and plant. In plants, dsRNAs targeted to CpG islands within a promoter can also induce RNA-directed DNA methylation; however, it remains unclear whether gene silencing mediated by DNA methylation can be induced by dsRNAs in mammalian cells. Here, we demonstrate that short interfering RNAs (siRNAs; 21-25-nucleotide RNA molecules) induce DNA methylation and histone H3 methylation in human cells. Synthetic siRNAs targeted to CpG islands of an E-cadherin promoter induced significant DNA methylation and histone H3 lysine 9 methylation in both MCF-7 and normal mammary epithelial cells. As a result, these siRNAs repressed expression of the E-cadherin gene at the transcriptional level. In addition, disrupting the expression of either one of two DNA methyltransferases (DNMT1 or DNMT3B) by specific siRNAs abolished the siRNA-mediated methylation of DNA. Moreover, vector-based siRNAs targeted to the erbB2 (also known as HER2) promoter also induced DNA methylation in MCF-7 cells. Thus, siRNAs targeted to CpG islands within the promoter of a specific gene can induce transcriptional gene silencing by means of DNA-methyltransferase-dependent methylation of DNA in human cells, and might have potential as a new type of gene therapeutic agent.
Department of Chemistry and Biotechnology, School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan. kawasaki@chembio.t.u-tokyo.ac.jp
Hiroaki
H
Kawasaki
Kazunari
K
Taira
D015820:Cadherins / Q000235:genetics; D002460:Cell Line; D045744:Cell Line, Tumor; D018899:CpG Islands / Q000235:genetics; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000235:genetics / Q000378:metabolism; D019175:DNA Methylation; D020868:Gene Silencing; D018734:Genes, erbB-2 / Q000235:genetics; D015316:Genetic Therapy / Q000379:methods; D006657:Histones / Q000378:metabolism; D006801:Humans; D008745:Methylation; D011401:Promoter Regions, Genetic / Q000235:genetics; D012330:RNA, Double-Stranded / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D000097584:DNA Methyltransferase 3B
D015820:Cadherins; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; D006657:Histones; D012330:RNA, Double-Stranded; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D000097584:DNA Methyltransferase 3B; C000619380:DNMT1 protein, human