This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow An author's correction has been published
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Araujo, F. D.
Right arrow Articles by Zannis-Hadjopoulos, M.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Araujo, F. D.
Right arrow Articles by Zannis-Hadjopoulos, M.

 Previous Article  |  Next Article 

Mol Cell Biol, June 1998, p. 3475-3482, Vol. 18, No. 6
0270-7306/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Concurrent Replication and Methylation at Mammalian Origins of Replication

Felipe D. Araujo,1,2 J. David Knox,3 Moshe Szyf,3,* Gerald B. Price,1 and Maria Zannis-Hadjopoulos1,2

McGill Cancer Centre,1 Department of Biochemistry,2 and Department of Pharmacology and Therapeutics,3 McGill University, Montreal, Quebec, Canada H3G 1Y6

Received 30 June 1997/Returned for modification 16 December 1997/Accepted 12 March 1998

Observations made with Escherichia coli have suggested that a lag between replication and methylation regulates initiation of replication. To address the question of whether a similar mechanism operates in mammalian cells, we have determined the temporal relationship between initiation of replication and methylation in mammalian cells both at a comprehensive level and at specific sites. First, newly synthesized DNA containing origins of replication was isolated from primate-transformed and primary cell lines (HeLa cells, primary human fibroblasts, African green monkey kidney fibroblasts [CV-1], and primary African green monkey kidney cells) by the nascent-strand extrusion method followed by sucrose gradient sedimentation. By a modified nearest-neighbor analysis, the levels of cytosine methylation residing in all four possible dinucleotide sequences of both nascent and genomic DNAs were determined. The levels of cytosine methylation observed in the nascent and genomic DNAs were equivalent, suggesting that DNA replication and methylation are concomitant events. Okazaki fragments were also demonstrated to be methylated, suggesting that the rapid kinetics of methylation is a feature of both the leading and the lagging strands of nascent DNA. However, in contrast to previous observations, neither nascent nor genomic DNA contained detectable levels of methylated cytosines at dinucleotide contexts other than CpG (i.e., CpA, CpC, and CpT are not methylated). The nearest-neighbor analysis also shows that cancer cell lines are hypermethylated in both nascent and genomic DNAs relative to the primary cell lines. The extent of methylation in nascent and genomic DNAs at specific sites was determined as well by bisulfite mapping of CpG sites at the lamin B2, c-myc, and beta -globin origins of replication. The methylation patterns of genomic and nascent clones are the same, confirming the hypothesis that methylation occurs concurrently with replication. Interestingly, the c-myc origin was found to be unmethylated in all clones tested. These results show that, like genes, different origins of replication exhibit different patterns of methylation. In summary, our results demonstrate tight coordination of DNA methylation and replication, which is consistent with recent observations showing that DNA methyltransferase is associated with proliferating cell nuclear antigen in the replication fork.

* Corresponding author. Mailing address: Department of Pharmacology, McGill University, 3655 Drummond St., Montreal, PQ H3G1Y6 Canada. Phone: (514) 398-7107. Fax: (514) 398-6690. E-mail: M52YF{at}Pharma.McGill.CA.

Mol Cell Biol, June 1998, p. 3475-3482, Vol. 18, No. 6
0270-7306/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:

  • Hermann, A., Goyal, R., Jeltsch, A. (2004). The Dnmt1 DNA-(cytosine-C5)-methyltransferase Methylates DNA Processively with High Preference for Hemimethylated Target Sites. J. Biol. Chem. 279: 48350-48359 [Abstract] [Full Text]  
  • Okuwaki, M., Verreault, A. (2004). Maintenance DNA Methylation of Nucleosome Core Particles. J. Biol. Chem. 279: 2904-2912 [Abstract] [Full Text]  
  • Ushijima, T., Watanabe, N., Okochi, E., Kaneda, A., Sugimura, T., Miyamoto, K. (2003). Fidelity of the Methylation Pattern and Its Variation in the Genome. Genome Res 13: 868-874 [Abstract] [Full Text]  
  • Milutinovic, S., Zhuang, Q., Niveleau, A., Szyf, M. (2003). Epigenomic Stress Response. KNOCKDOWN OF DNA METHYLTRANSFERASE 1 TRIGGERS AN INTRA-S-PHASE ARREST OF DNA REPLICATION AND INDUCTION OF STRESS RESPONSE GENES. J. Biol. Chem. 278: 14985-14995 [Abstract] [Full Text]  
  • Milutinovic, S., Zhuang, Q., Szyf, M. (2002). Proliferating Cell Nuclear Antigen Associates with Histone Deacetylase Activity, Integrating DNA Replication and Chromatin Modification. J. Biol. Chem. 277: 20974-20978 [Abstract] [Full Text]  
  • Liang, G., Chan, M. F., Tomigahara, Y., Tsai, Y. C., Gonzales, F. A., Li, E., Laird, P. W., Jones, P. A. (2002). Cooperativity between DNA Methyltransferases in the Maintenance Methylation of Repetitive Elements. Mol. Cell. Biol. 22: 480-491 [Abstract] [Full Text]  
  • Meehan, R. R., Pennings, S., Stancheva, I. (2001). Lashings of DNA methylation, forkfuls of chromatin remodeling. Genes Dev. 15: 3231-3236 [Full Text]  
  • Singal, R., vanWert, J. M. (2001). De novo methylation of an embryonic globin gene during normal development is strand specific and spreads from the proximal transcribed region. Blood 98: 3441-3446 [Abstract] [Full Text]  
  • Lin, X., Tascilar, M., Lee, W.-H., Vles, W. J., Lee, B. H., Veeraswamy, R., Asgari, K., Freije, D., van Rees, B., Gage, W. R., Bova, G. S., Isaacs, W. B., Brooks, J. D., DeWeese, T. L., De Marzo, A. M., Nelson, W. G. (2001). GSTP1 CpG Island Hypermethylation Is Responsible for the Absence of GSTP1 Expression in Human Prostate Cancer Cells. Am. J. Pathol. 159: 1815-1826 [Abstract] [Full Text]  
  • Lin, C.-H., Hsieh, S.-Y., Sheen, I-S., Lee, W.-C., Chen, T.-C., Shyu, W.-C., Liaw, Y.-F. (2001). Genome-wide Hypomethylation in Hepatocellular Carcinogenesis. Cancer Res. 61: 4238-4243 [Abstract] [Full Text]  
  • Milutinovic, S., Knox, J. D., Szyf, M. (2000). DNA Methyltransferase Inhibition Induces the Transcription of the Tumor Suppressor p21WAF1/CIP1/sdi1. J. Biol. Chem. 275: 6353-6359 [Abstract] [Full Text]  
  • Radomski, N., Kaufmann, C., Dreyer, C. (1999). Nuclear Accumulation of S-Adenosylhomocysteine Hydrolase in Transcriptionally Active Cells during Development of Xenopus laevis. Mol. Biol. Cell 10: 4283-4298 [Abstract] [Full Text]  
  • Bacolla, A., Pradhan, S., Roberts, R. J., Wells, R. D. (1999). Recombinant Human DNA (Cytosine-5) Methyltransferase. II. STEADY-STATE KINETICS REVEAL ALLOSTERIC ACTIVATION BY METHYLATED DNA. J. Biol. Chem. 274: 33011-33019 [Abstract] [Full Text]  
  • Rein, T., Kobayashi, T., Malott, M., Leffak, M., DePamphilis, M. L. (1999). DNA Methylation at Mammalian Replication Origins. J. Biol. Chem. 274: 25792-25800 [Abstract] [Full Text]  
  • De Marzo, A. M., Marchi, V. L., Yang, E. S., Veeraswamy, R., Lin, X., Nelson, W. G. (1999). Abnormal Regulation of DNA Methyltransferase Expression during Colorectal Carcinogenesis. Cancer Res. 59: 3855-3860 [Abstract] [Full Text]  
  • Singal, R., Ginder, G. D. (1999). DNA Methylation. Blood 93: 4059-4070 [Full Text]  
  • Araujo, F. D., Knox, J. D., Ramchandani, S., Pelletier, R., Bigey, P., Price, G., Szyf, M., Zannis-Hadjopoulos, M. (1999). Identification of Initiation Sites for DNA Replication in the Human dnmt1 (DNA-methyltransferase) Locus. J. Biol. Chem. 274: 9335-9341 [Abstract] [Full Text]  
  • Bigey, P., Knox, J. D., Croteau, S., Bhattacharya, S. K., Theberge, J., Szyf, M. (1999). Modified Oligonucleotides as Bona Fide Antagonists of Proteins Interacting with DNA. HAIRPIN ANTAGONISTS OF THE HUMAN DNA METHYLTRANSFERASE. J. Biol. Chem. 274: 4594-4606 [Abstract] [Full Text]  
  • Knox, J. D., Araujo, F. D., Bigey, P., Slack, A. D., Price, G. B., Zannis-Hadjopoulos, M., Szyf, M. (2000). Inhibition of DNA Methyltransferase Inhibits DNA Replication. J. Biol. Chem. 275: 17986-17990 [Abstract] [Full Text]  
  • Detich, N., Ramchandani, S., Szyf, M. (2001). A Conserved 3'-Untranslated Element Mediates Growth Regulation of DNA Methyltransferase 1 and Inhibits Its Transforming Activity. J. Biol. Chem. 276: 24881-24890 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»