Double RNA interference of DNMT3b and DNMT1 enhances DNA demethylation and gene reactivation

Yu Wei Leu, Farahnaz Rahmatpanah, Huidong Shi, Susan H. Wei, Joseph C. Liu, Pearlly S. Yan, Hui-ming Huang

Research output: Contribution to journalArticle

127 Citations (Scopus)

Abstract

Small interfering RNAs (siRNAs) are newly identified molecules shown to silence genes via targeted mRNA degradation. In this study, we used specific siRNAs as a tool to probe the relationship between two DNA methyltransferase genes, DNMT3b and DNMT1, in the maintenance of DNA methylation patterns in the genome. Levels of DNMT3b or DNMT1 mRNAs and proteins were markedly decreased (up to 80%) on transfecting these siRNAs into the ovarian cancer cell line CP70. The resulting RNA interference showed differential effects on DNA demethylation and gene reactivation in the treated cells. The DNMT1 siRNA treatment led to a partial removal of DNA methylation from three inactive promoter CpG islands, TWIST, RASSF1A, and HIN-1, and restored the expression of these genes. This epigenetic alteration appeared less effective in cells transfected with DNMT3b siRNA. However, the combined treatment of DNMT3b and DNMT1 siRNAs greatly enhanced this demethylation effect, producing 7-15-fold increases in their expression. We also used a microarray approach to examine this RNA interference on 8640 CpG island loci in CP70 cells. The combined siRNA treatment had a greater demethylation effect on 241 methylated loci and selected repetitive sequences than that of the single treatment. Our data thus suggest that whereas DNMT1 plays a key role in methylation maintenance, DNMT3b may act as an accessory to support the function in CP70 cells. This study also shows that siRNA is a powerful tool for interrogating the mechanisms of DNA methylation in normal and pathological genomes.

Original languageEnglish (US)
Pages (from-to)6110-6115
Number of pages6
JournalCancer Research
Volume63
Issue number19
StatePublished - Oct 1 2003
Externally publishedYes

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RNA Interference
Small Interfering RNA
DNA
Genes
DNA Methylation
CpG Islands
Maintenance
Genome
Nucleic Acid Repetitive Sequences
RNA Stability
Methyltransferases
Epigenomics
Ovarian Neoplasms
Methylation
Gene Expression
Cell Line
Messenger RNA

ASJC Scopus subject areas

  • Cancer Research
  • Oncology

Cite this

Leu, Y. W., Rahmatpanah, F., Shi, H., Wei, S. H., Liu, J. C., Yan, P. S., & Huang, H. (2003). Double RNA interference of DNMT3b and DNMT1 enhances DNA demethylation and gene reactivation. Cancer Research, 63(19), 6110-6115.

Double RNA interference of DNMT3b and DNMT1 enhances DNA demethylation and gene reactivation. / Leu, Yu Wei; Rahmatpanah, Farahnaz; Shi, Huidong; Wei, Susan H.; Liu, Joseph C.; Yan, Pearlly S.; Huang, Hui-ming.

In: Cancer Research, Vol. 63, No. 19, 01.10.2003, p. 6110-6115.

Research output: Contribution to journalArticle

Leu, YW, Rahmatpanah, F, Shi, H, Wei, SH, Liu, JC, Yan, PS & Huang, H 2003, 'Double RNA interference of DNMT3b and DNMT1 enhances DNA demethylation and gene reactivation', Cancer Research, vol. 63, no. 19, pp. 6110-6115.
Leu YW, Rahmatpanah F, Shi H, Wei SH, Liu JC, Yan PS et al. Double RNA interference of DNMT3b and DNMT1 enhances DNA demethylation and gene reactivation. Cancer Research. 2003 Oct 1;63(19):6110-6115.
Leu, Yu Wei ; Rahmatpanah, Farahnaz ; Shi, Huidong ; Wei, Susan H. ; Liu, Joseph C. ; Yan, Pearlly S. ; Huang, Hui-ming. / Double RNA interference of DNMT3b and DNMT1 enhances DNA demethylation and gene reactivation. In: Cancer Research. 2003 ; Vol. 63, No. 19. pp. 6110-6115.
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