Deletions of a differentially methylated CpG island at the SNRPN gene define a putative imprinting control region

To determine the molecular basis of Prader-Willi syndrome (PWS) and Angelman syndrome (AS), we have isolated new transcripts from chromosome 15q11−q13. Two novel transcripts located within 300 kilobases telomeric to the small nuclear ribonucleoprotein-associated polypeptide N gene (SNRPN) were paternally expressed in cultured cells, along with SNRPN, defining a large imprinted transcriptional domain. In three PWS patients (two sibs), small deletions remove a differentially methylated CpG island containing a newly described 5' exon of SNRPN, and cause loss of expression for the three imprinted transcripts and altered methylation over hundreds of kilobases. The smallest PWS deletion is familial and asymptomatic with maternal transmission. Our data imply the presence of a paternal imprinting control region near exon .

1. Ledbetter, D.H. et al. Deletions of chromosome 15 as a cause of the Prader−Willi syndrome. New Engl. J. Med. 304, 325−329 (1981). | PubMed  | ISI | ChemPort |
2. Knoll, J.H.M. et al. Angelman and Prader-Willi syndromes share a common chromosome 15 deletion but differ in parental origin of the deletion. Am. J. med. Genet. 32, 285−290 (1989). | PubMed  | ISI | ChemPort |
3. Nicholls, R.D., Knoll, J.H.M., Butler, M.G., Karam, S. & Lalande, M. Genetic imprinting suggested by maternal heterodisomy in non-deletion Prader-Willi syndrome. Nature 342, 281−285 (1989). | Article | PubMed  | ISI | ChemPort |
4. Malcolm, S. et al. Uniparental paternal disomy in Angelman's syndrome. Lancet 337, 694−697 (1991). | Article | PubMed  | ISI | ChemPort |
5. Holm, V.A. et al. Prader-Willi syndrome: consensus diagnostic criteria. Pediatrics 91, 398−402 (1993). | PubMed  | ISI | ChemPort |
6. Robinson, W.P. et al. Molecular, cytogenetic, and clinical investigations of Prader-Willi syndrome patients. Am. J. hum. Genet. 49, 1219−1234 (1991). | PubMed  | ISI | ChemPort |
7. Mutirangura, A. et al. A complete YAC contig of the Prader−Willi/Angelman chromosome region (15q11−q13) and refined localization of the SNRPN gene. Genomics 18, 546−552 (1993). | PubMed  | ISI | ChemPort |
8. Reis, A. et al. Imprinting mutations suggested by abnormal DNA methylation patterns in familial Angelman and Prader−Willi syndromes. Am. J. hum. Genet. 54, 741−747 (1994). | PubMed  | ISI | ChemPort |
9. Mascari, M.J. et al. The frequency of uniparental disomy in Prader−Willi syndrome. Implications for molecular diagnosis. New Engl. J. Med. 326, 1599−1607 (1992). | PubMed  | ISI | ChemPort |
10. Meijers-Heijboer, E.J. et al. Linkage analysis with chromosome 15q11−13 markers shows genomic imprinting in familial Angelman syndrome. J. med. Genet. 29, 853−857 (1992). | PubMed  | ChemPort |
11. Clayton-Smith, J. et al. Further evidence for dominant inheritance at the chromosome 15q11−13 locus in familial Angelman syndrome. Am J. med. Genet. 44, 256−260 (1992). | PubMed  | ChemPort |
12. Wagstaff, J. et al. Maternal but not paternal transmission of 15q11−13-linked nondeletion Angelman syndrome leads to phenotypic expression. Nature Genet. 1, 291−294 (1992). | Article | PubMed  | ISI | ChemPort |
13. Kuwano, A. et al. Molecular dissection of the Prader-Willi/Angelman syndrome region (15q11−13) by YAC cloning and FISH analysis. Hum. molec. Genet. 1, 417−425 (1992). | PubMed  | ChemPort |
14. Reis, A. et al. Exclusion of the GABA-receptor b3 subunit gene as the Angelman's syndrome gene. Lancet 341, 122−123 (1993). | Article | PubMed  | ISI | ChemPort |
15. Hamabe, J. et al. DNA deletion and its parental origin in Angelman syndrome patients. Am. J. med. Genet. 41, 64−68 (1991). | PubMed  | ISI | ChemPort |
16. Saitoh, S. et al. Familial Angelman syndrome caused by imprinted submicroscopic deletion encompassing GABA_A receptor b3 subunit gene. Lancet 339, 366−367 (1992). | Article | PubMed  | ISI | ChemPort |
17. Özçelik, T. et al. Small nuclear ribonucleoprotein polypeptide N (SNRPN), an expressed gene in the Prader-Willi syndrome critical region. Nature Genet. 2, 265−269 (1992). | Article | PubMed  | ISI | ChemPort |
18. Leff, S.E. et al. Maternal imprinting of the mouse Snrpn gene and conserved linkage homology with the human Prader-Willi syndrome region. Nature Genet. 2, 259−264 (1992). | Article | PubMed  | ISI | ChemPort |
19. Cattanach, B.M. et al. A candidate mouse model for Prader-Willi syndrome which shows an absence of Snrpn expression. Nature Genet. 2, 270−274 (1992). | Article | PubMed  | ISI | ChemPort |
20. Nakao, M. et al. Imprinting analysis of three genes in the Prader−Willi/ Angelman region: SNRPN, E6-associated protein, and PAR-2 (D15S225E). Hum. molec. Genet. 3, 309−315 (1994). | PubMed  | ISI | ChemPort |
21. Glenn, C.C., Porter, K.A., Jong, M.T.C., Nicholls, R.D. & Driscoll, D.J. Functional imprinting and epigenetic modification of the human SNRPN gene. Hum. molec. Genet. 2, 2001−2005 (1993). | PubMed  | ISI | ChemPort |
22. Reed, M.L. & Leff, S.E. Maternal imprinting of human SNRPN, a gene deleted in the Prader-Willi syndrome. Nature Genet. 6, 163−167 (1994). | Article | PubMed  | ISI | ChemPort |
23. Dittrich, B. et al. Molecular diagnosis of the Prader−Willi and Angelman syndromes by detection of parent-of-origin specific DNA methylation in 15q11−13. Hum. Genet. 90, 313−315 (1992). | PubMed  | ISI | ChemPort |
24. Driscoll, D.J. et al. A DNA methylation imprint, determined by the sex of the parent, distinguishes the Angelman and Prader−Willi syndromes. Genomics 13, 917−924 (1992). | PubMed  | ISI | ChemPort |
25. Glenn, C.C. et al. Modification of 15q11−q13 DNA methylation imprints in unique Angelman and Prader-Willi patients. Hum. molec. Genet. 2, 1377−1382 (1993). | PubMed  | ISI | ChemPort |
26. Nicholls, R.D. New insights reveal complex mechanisms involved in genomic imprinting. Am. J. hum. Genet. 54, 733−740 (1994). | PubMed  | ISI | ChemPort |
27. Huibregtse, J.M., Scheffner, M. & Howley, P.M. Cloning and expression of the cDNA for E6-AP, a protein that mediates the interaction of the human papillomavirus E6 oncoprotein with p53. Molec. cell. Biol. 13, 775−784 (1993). | PubMed  | ISI | ChemPort |
28. Sutcliffe, J.S., Zhang, F., Caskey, C.T., Nelson, D.L. & Warren, S.T. PCR amplification and analysis of yeast artificial chromosomes. Genomics 13, 1303−1306 (1992). | PubMed  | ISI | ChemPort |
29. McAllister, G., Amara, S.G. & Lemer, M.R. Tissue-specific expression and cDNA cloning of small nuclear ribonucleoprotein-associated polypeptide N. Proc. natn. Acad. Sci. U.S.A. 85, 5296−5300 (1988). | ChemPort |
30. Örstavik, K.H. et al. Prader−Willi syndrome in a brother and sister without cytogenetic or detectable molecular genetic abnormality at chromosome 15q11q13. Am. J. med. Genet. 44, 534−538 (1992). | PubMed  | ISI |
31. Pettigrew, A.L., Gollin, S.M., Greenberg, F., Riccardi, V.M. & Ledbetter, D.H. Duplication of proximal 15q as a cause of Prader−Willi syndrome. Am. J. med. Genet. 28, 791−802 (1987). | PubMed  | ISI | ChemPort |
32. Schmauss, C., Brines, M.L. & Lemer, M.R. The gene encoding the small nuclear ribonucleoprotein-associated protein N is expressed at high levels in neurons. J. biol. Chem. 267, 8521−8529 (1992). | PubMed  | ISI | ChemPort |
33. Schmauss, C., McAllister, G., Ohosone, Y., Hardin, J.A. & Lerner, M.R. A comparison of snRNP-associated Sm-autoantigens: human N, rat N and human B/B'. Nucl. Acids Res. 17, 1733−1743 (1989). | PubMed  | ISI | ChemPort |
34. Rokeach, L.A., Jannatipour, M., Haselby, J.A. & Hoch, S.O. Primary structure of a human small nuclear ribonucleoprotein polypeptide as deduced by cDNA analysis. J. biol. Chem. 264, 5024−5030 (1989). | PubMed  | ISI | ChemPort |
35. Brown, C.J. et al. The human XIST gene: analysis of a 17 kb inactive X-specific RNA that contains conserved repeats and is highly localized within the nucleus. Cell 71, 527−542 (1992). | Article | PubMed  | ISI | ChemPort |
36. Zemel, S., Bartolomei, M.S. & Tilghman, S.M. Physical linkage of two mammalian imprinted genes, H19 and insulin-like growth factor 2. Nature Genet. 2, 61−65 (1992). | Article | PubMed  | ISI | ChemPort |
37. Bartolomei, M.S. & Tilghman, S.M. Parental imprinting of mouse chromosome 7. Sem. Devi. Biol. 3, 107−117 (1992).
38. Bartolomei, M.S., Webber, A.L., Brunkow, M.E. & Tilghman, S.M. Epigenetic mechanisms underlying the imprinting of the mouse H19 gene. Genes Dev. 7, 1663−1673 (1993). | PubMed  | ISI | ChemPort |
39. Li, E., Beard, C. & Jaenisch, R. Role for DNA methylation in genomic imprinting. Nature 366, 362−365 (1993). | Article | PubMed  | ISI | ChemPort |
40. Southern, E.M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J. molec. Biol. 98, 503−517 (1975). | PubMed  | ISI | ChemPort |
41. Feinberg, A.P. & Vogelstein, B. A technique for radiolabelling DNA restriction fragments to high specific activity. Anal. Biochem. 132, 6−13 (1983). | PubMed  | ISI | ChemPort |
42. Sealy, P.G., Whittaker, P.A. & Southern, E.M. Removal of repeated sequences from hybridization probes. Nucl. Acids Res. 13, 1905−1922 (1985). | PubMed  | ISI | ChemPort |
43. Albertsen, H.M. et al. Construction and characterization of a yeast artificial chromosome library containing seven haploid human genome equivalents. Proc. natn. Acad. Sci. U.S.A. 87, 4256−4260 (1990). | ChemPort |
44. Anand, R., Villasante, A. & Tyler-Smith, C. Construction of yeast artificial chromosome libraries with large inserts using fractionation by pulsed-field gel electrophoresis. Nucl. Acids Res. 17, 3425−3433 (1989). | PubMed  | ISI | ChemPort |
45. Smith, C.L., Klco, S.R. & Cantor, C.R. in Genome Analysis: A Practical Approach (ed. Davies, K.E. ) 41−72 (IRL Press, Oxford, 1988).
46. Chomczynski, P. & Sacchi, N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chioroform extraction. Anal. Biochem. 162, 156−159 (1987). | Article | PubMed  | ISI | ChemPort |
47. Kuwano, A., Ledbetter, S.A., Dobyns, W.B., Emanuel, B.S. & Ledbetter, D.H. Detection of deletions and cryptic translocations in Miller-Dieker syndrome by in situ hybridization. Am. J. hum. Genet. 49, 707−714 (1991). | PubMed  | ISI | ChemPort |
48. Zoghbi, H.Y. et al. Assignment of autosomal dominant spinocerebellar ataxia (SCA1) centromeric to the HLA region on the short arm of chromosome 6, using multilocus linkage analysis. Am. J. hum. Genet. 44, 255−263 (1989). | PubMed  | ISI | ChemPort |

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