Abstract
An expansion of a CTG repeat at the DM1 locus causes myotonic dystrophy (DM) by altering the expression of the two adjacent genes, DMPK and SIX5, and through a toxic effect of the repeat-containing RNA. Here we identify two CTCF-binding sites that flank the CTG repeat and form an insulator element between DMPK and SIX5. Methylation of these sites prevents binding of CTCF, indicating that the DM1 locus methylation in congenital DM would disrupt insulator function. Furthermore, CTCF-binding sites are associated with CTG/CAG repeats at several other loci. We suggest a general role for CTG/CAG repeats as components of insulator elements at multiple sites in the human genome.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 12 print issues and online access
$209.00 per year
only $17.42 per issue
Rent or buy this article
Prices vary by article type
from$1.95
to$39.95
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Brook, J.D.M.C. et al. Molecular basis of myotonic dystrophy: expansion of a trinucleotide (CTG) repeat at the 3′ end of a transcript encoding a protein kinase family member. Cell 68, 799–808 (1992).
Harley, H.G. et al. Expansion of an unstable DNA region and phenotypic variation in myotonic dystrophy. Nature 355, 545–546 (1992).
Mahadevan, M. et al. Myotonic dystrophy mutation: an unstable CTG repeat in the 3′ untranslated region of the gene. Science 255, 1253–1255 (1992).
Fu, Y.H. et al. An unstable triplet repeat in a gene related to myotonic muscular dystrophy. Science 255, 1256–1258 (1992).
Tsilfidis, C., MacKenzie, A.E., Mettler, G., Barcelo, J. & Korneluk, R.G. Correlation between CTG trinucleotide repeat length and frequency of severe congenital myotonic dystrophy. Nature Genet. 1, 192–195 (1992).
Steinbach, P., Glaser, D., Walther, V., Wolf, M. & Schwemmle, S. The DMPK gene of severely affected myotonic dystrophy patients is hypermethylated proximal to the largely expanded CTG repeat. Am. J. Hum. Genet. 62, 278–285 (1998).
Tapscott, S.J. Deconstructing myotonic dystrophy. Science 289, 1701–1702 (2000).
Novelli, G.G. et al. Failure in detecting mRNA transcripts from the mutated allele in myotonic dystrophy muscle. Biochem. Mol. Biol. Int. 29, 291–297 (1993).
Fu, Y.H. et al. Decreased expression of myotonin-protein kinase messenger RNA and protein in adult form of myotonic dystrophy. Science 260, 235–238 (1993).
Taneja, K.L., McCurrach, M., Schalling, M., Housman, D. & Singer, R.H. Foci of trinucleotide repeat transcripts in nuclei of myotonic dystrophy cells and tissues. J. Cell Biol. 128, 995–1002 (1995).
Krahe, R. et al. Effect of myotonic dystrophy trinucleotide repeat expansion on DMPK transcription and processing. Genomics 28, 1–14 (1995).
Berul, C.I. et al. DMPK dosage alterations result in atrioventricular conduction abnormalities in a mouse myotonic dystrophy model. J. Clin. Invest. 103, R1–R7 (1999).
Klesert, T.R., Otten, A.D., Bird, T,D. & Tapscott, S.J. Trinucleotide repeat expansion at the myotonic dystrophy locus reduces expression of DMAHP. Nature Genet. 16, 402–406 (1997).
Thornton, C.A., Wymer, J.P., Simmons, Z., McClain, C. & Moxley III, R.T. Expansion of the myotonic dystrophy CTG repeat reduces expression of the flanking DMAHP gene. Nature Genet. 16, 407–409 (1997).
Klesert, T.R. et al. Mice deficient in Six5 develop cataracts: implications for myotonic dystrophy. Nature Genet. 25, 105–109 (2000).
Sarkar, P.S. et al. Heterozygous loss of Six5 in mice is sufficient to cause ocular cataracts. Nature Genet. 25, 110–114 (2000).
Mankodi, A. et al. Myotonic dystrophy in transgenic mice expressing an expanded CUG repeat. Science 289, 1769–1773 (2000).
Sabouri, L.A. et al. Effect of the myotonic dystrophy (DM) mutation on mRNA levels of the DM gene. Nature Genet. 4, 233–238 (1993).
Alwazzan, M., Hamshere, M.G., Lennon, G.G. & Brook, J.D. Six transcripts map within 200 kilobases of the myotonic dystrophy expanded repeat. Mamm. Genome 9, 485–487 (1998).
Bell, A.C., West, A.G. & Felsenfeld, G. Gene Regulation: insulators and boundaries: versatile regulatory elements in the eukaryotic genome. Science 291, 447–450 (2001).
Bell, A.C., West, A.G. & Felsenfeld, G. The protein CTCF is required for the enhancer blocking activity of vertebrate insulators. Cell 98, 387–396 (1999).
Bell, A.C. & Felsenfeld, G. Methylation of a CTCF-dependent boundary controls imprinted expression of the Igf2 gene. Nature 405, 482–485 (2000).
Hark, A.T. et al. CTCF mediates methylation-sensitive enhancer-blocking activity at the H19/Igf2 locus. Nature 405, 486–489 (2000).
Kanduri, C. et al. Functional association of CTCF with the insulator upstream of the H19 gene is parent of origin-specific and methylation-sensitive. Curr. Biol. 10, 853–856 (2000).
Mirkovitch, J., Mirault, M.E. & Laemmli, U.K. Organization of the higher-order chromatin loop: specific DNA attachment sites on nuclear scaffold. Cell 39, 223–232 (1984).
Filippova, G. et al. An exceptionally conserved transcriptional repressor, CTCF, employs different combinations of zinc fingers to bind diverged promoter sequences of avian and mammalian c-myc oncogenes. Mol. Cell. Biol. 16, 2802–2813 (1996).
Burcin, M. et al. Negative protein 1, which is required for function of the chicken lysozyme gene silencer in conjunction with hormone receptors, is identical to the multivalent zinc finger repressor CTCF. Mol. Cell. Biol. 17, 1281–1288 (1997).
Vostrov, A. & Quitschke, W. The zinc finger protein CTCF binds to the APB-beta domain of the amyloid beta-protein precursor promoter: evidence for a role in transcriptional activation. J. Biol. Chem. 272, 33353–33359 (1997).
Filippova, G.N. et al. A widely expressed transcription factor with multiple DNA sequence specificity, CTCF, is localized at chromosome segment 16q22.1 within one of the smallest regions of overlap for common deletions in breast and prostate cancers. Genes Chrom. Cancer 22, 26–36 (1998).
Wang, Y-H., Amirhaeri, S., Kang, S., Wells, R.D. & Griffith, J.D. Preferential nucleosome assembly at DNA triplet repeats from the myotonic dystrophy gene. Science 265, 669–671 (1994).
Godde, J.S. & Wolffe, A.P. Nucleosome assembly on CTG triplet repeats. J. Biol. Chem. 271, 15222–15229 (1996).
Wang, Y. & Griffith, J. Expanded CTG triplet blocks from the myotonic dystrophy gene create the strongest known natural nucleosome positioning elements. Genomics 25, 570–573 (1995).
Chung, J.H., Whiteley, M. & Felsenfeld, G. A 5′ element of the chicken beta-globin domain serves as an insulator in human erythroid cells and protects against position effect in Drosophila. Cell 74, 505–514 (1993).
Lobanenkov, V.V., Nicolas, R.H., Plumb, M.A., Wright, C.A. & Goodwin, G.H. Sequence-specific DNA-binding proteins which interact with (G+C)-rich sequences flanking the chicken c-myc gene. Eur. J. Biochem. 159, 181–188 (1986).
Lobanenkov, V.V. et al. A novel sequence-specific DNA binding protein which interacts with three regularly spaced direct repeats of the CCCTC-motif in the 5′-flanking sequence of the chicken c-myc gene. Oncogene 5, 1743–1753 (1990).
Klenova, E.M. et al. CTCF, a conserved nuclear factor required for optimal transcriptional activity of the chicken cmyc gene, is an 11-Zn-finger protein differentially expressed in multiple forms. Mol. Cell. Biol. 13, 7612–7624 (1993).
Awad, T.A. et al. Negative transcriptional regulation mediated by thyroid hormone response element 144 requires binding of the multivalent factor CTCF to a novel DNA sequence. J. Biol. Chem. 274, 27092–27098 (1999).
Wolffe, A.P. Imprinting insulation. Curr. Biol. 10, R463–465 (2000).
Reik, W. & Murrell, A. Silence across the border. Nature 405, 408–409 (2000).
Reddy, P.S. & Housman, D.E. The complex pathology of trinucleotide repeats. Curr. Opin. Cell. Biol. 9, 364–372 (1997).
Kanduri, C. et al. The 5′-flank of the murine H19 gene in an unusual chromatin conformation unidirectionally blocks enhancer-promoter communication. Curr. Biol. 10, 449–457 (2000).
Wang, Y.H., Gellibolian, R., Shimizu, M., Wells, R.D. & Griffith, J. Long CCG triplet repeat blocks exclude nucleosomes: a possible mechanism for the nature of fragile sites in chromosomes. J. Mol. Biol. 263, 511–516 (1996).
Gerber, A.N., Klesert, T.R., Bergstrom, D.A. & Tapscott, S.J. Two domains of MyoD mediate transcriptional activation of genes in repressive chromatin: a mechanism for lineage determination in myogenesis. Genes Dev. 11, 436–450 (1997).
Mahadevan, M.S. et al. Structure and genomic sequence of the myotonic dystrophy (DM kinase) gene. Hum. Mol. Genet. 2, 299–304 (1993).
Boucher, C.A. et al. A novel homeodomain-encoding gene is associated with a large CpG island interrupted by the myotonic dystrophy unstable (CTG)n repeat. Hum. Mol. Genet. 4, 1919–1925 (1995).
Acknowledgements
This work was supported by National Institutes of Health, National Institute of Arthritis and Musculoskeletal and Skin Disease R01-AR45203 (S.J.T.) and National Institutes of Health R01-CA68360 (G.N.F.). We thank P. Rollini and K. Fournier for help with MAR assays, L. Ashworth at the Lawrence Livermore National Laboratory Genome Center for her help in compiling a contiguous sequence for the DM1 locus, J.D. Brook and M.G. Hamshere for cosmids, A.C. Bell, G. Felsenfeld, and C.A. Thornton for plasmids and P. Neiman, S. Collins, M. Groudine and B. Trask for critical comments on the manuscript.
Author information
Authors and Affiliations
Corresponding authors
Rights and permissions
About this article
Cite this article
Filippova, G., Thienes, C., Penn, B. et al. CTCF-binding sites flank CTG/CAG repeats and form a methylation-sensitive insulator at the DM1 locus. Nat Genet 28, 335–343 (2001). https://doi.org/10.1038/ng570
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/ng570
This article is cited by
-
CGGBP1-regulated cytosine methylation at CTCF-binding motifs resists stochasticity
BMC Genetics (2020)
-
Targeting CTCFL/BORIS for the immunotherapy of cancer
Cancer Immunology, Immunotherapy (2018)
-
A screening system to identify transcription factors that induce binding site-directed DNA demethylation
Epigenetics & Chromatin (2017)
-
Contraction of fully expanded FMR1 alleles to the normal range: predisposing haplotype or rare events?
Journal of Human Genetics (2017)
-
Epigenetic drug discovery: breaking through the immune barrier
Nature Reviews Drug Discovery (2016)