Role of DNA methyltransferases in regulation of human ribosomal RNA gene transcription
Authors: Shoumei Bai
Kalpana Ghoshal
Samson T Jacob
Jharna Datta
Sarmila Majumder
David Spencer Smith
Journal: Journal of Biological Chemistry (2006)
Discipline: BIOLOGY & BIOCHEMISTRY
Keywords: ribosomal RNA genes; methylation; DNMTs; rDNA promoters; MBD2
Affiliations: Department of Molecular and Cellular Biochemistry, College of Medicine, Ohio State University, Columbus, Ohio 43210. Electronic address: ., Department of Molecular and Cellular Biochemistry, College of Medicine, Ohio State University, Columbus, Ohio 43210.

Abstract:  We have previously demonstrated that the expression of human ribosomal RNA genes (rDNA) in normal and cancer cells is differentially regulated by methylation of the promoter CpG islands. Furthermore, we showed that the methyl CpG-binding protein MBD2 plays a selective role in the methylation-mediated block in rDNA expression. Here, we analyzed the role of three functional mammalian DNA methyltransferases (DNMTs) in regulating the rDNA promoters activity. Immunofluorescence analysis and biochemical fractionation showed that all three DNMTs (DNMT1, DNMT3A, and DNMT3B) are associated with the inactive rDNA in the nucleolus. Although DNMTs associate with both methylated and unmethylated rDNA promoters, DNMT1 preferentially associates with the methylated genes. The rDNA primary transcript level was significantly elevated in DNMT1-/- or DNMT3B-/- human colon carcinoma (HCT116) cells. Southern blot analysis demonstrated a moderate level of rDNA promoter hypomethylation in DNMT1-/- cells and a dramatic loss of rDNA promoter methylation in double knockout cells. Transient overexpression of DNMT1 or DNMT3B suppressed the luciferase expression from both methylated and unmethylated pHrD-IRES-Luc, a reporter plasmid where the rDNA promoter drives luciferase expression. DNMT1-mediated suppression of the unmethylated promoter involves de novo methylation of the promoter, whereas histone deacetylase 2 cooperates with DNMT1 to inhibit the methylated rDNA promoter. Unlike DNMT1, both the wild type and catalytically inactive DNMT3B mutant can suppress rDNA promoter irrespective of its methylation status. DNMT3B-mediated suppression of the rDNA promoter also involves histone deacetylation. Treatment of HCT116 cells with Decitabine (a DNMT inhibitor) or trichostatin A (a histone deacetylase inhibitor) up-regulated endogenous rDNA expression. These inhibitors synergistically activated methylated pHrD-IRES-Luc, whereas they exhibited additive effects on the unmethylated promoter. These results demonstrate localization of DNMTs with the inactive rDNA in the nucleolus, the specific role of DNMT1 and DNMT3B in rDNA expression and the differential regulation of rDNA expression from the methylated and unmethylated rDNA promoters. 

Retracted: Yes

PubPeer comments:
null, 
 2018 retraction notice. http: www.jbc.org content 293 10 3591  VOLUME 281 (2006) PAGES 22062u201322072 This article has been withdrawn by the authors. In June 2017, the Journal questioned whether there were duplications in Figs. 1E and 6C and questioned the absence of data in one field of Fig. 5B. The original data and originally submitted figures were not available for evaluation. The authors are not convinced there are any duplications in Figs. 1E and 6C. Regarding Fig. 5B, the authors agree that the data for u201cVectoru201d was inadvertently omitted from the final figure. The authors were able to locate replicate data performed at the time of the original work, which they state shows an absence of expression in the Vector panel. The authors offered to publish a correction based on this repeat experiment and, alternatively, offered to repeat the experiment. However, the Journal declined both these offers, a decision with which the authors disagree. The authors stand by the reproducibility of the experimental data and the conclusions of the paper. The paper, with confirmatory data supporting the results, can be obtained by contacting the authors. u00a9 2018 by The American Society for Biochemistry and Molecular Biology, Inc. , 
 https: retractionwatch.com 2018 03 14 cancer-biologist-retracts-five-papers , 
null, 
null, 
 Penultimate author appears to currently be affiliated with University of Pittsburgh, and has the unfortunate distinction of appearing as coauthor of multiple papers with similar image concerns while at different institutions: https: pubpeer.com search?q=Shoumei+Bai  ,