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Conservation of the H19 noncoding RNA and H19-IGF2 imprinting mechanism in therians

Abstract

Comparisons between eutherians and marsupials suggest limited conservation of the molecular mechanisms that control genomic imprinting in mammals. We have studied the evolution of the imprinted IGF2-H19 locus in therians. Although marsupial orthologs of protein-coding exons were easily identified, the use of evolutionarily conserved regions and low-stringency Bl2seq comparisons was required to delineate a candidate H19 noncoding RNA sequence. The therian H19 orthologs show miR-675 and exon structure conservation, suggesting functional selection on both features. Transcription start site sequences and poly(A) signals are also conserved. As in eutherians, marsupial H19 is maternally expressed and paternal methylation upstream of the gene originates in the male germline, encompasses a CTCF insulator, and spreads somatically into the H19 gene. The conservation in all therians of the mechanism controlling imprinting of the IGF2-H19 locus suggests a sequential model of imprinting evolution.

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Figure 1: Characterization of the marsupial H19 gene.
Figure 2: Imprinted expression of the wallaby H19 and IGF2 genes.
Figure 3: The wallaby H19 differentially methylated domain is a CTCF insulator.
Figure 4: The marsupial H19 gene is a miRNA precursor.
Figure 5: Sequential model of the evolution of imprinting.

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Acknowledgements

The authors thank C. Edwards, A. Ferguson-Smith, E. Ivanova, G. Kelsey, W. Dean, S. Gaunt, J.M. Graves, K. Ishihara and M. Nakao for contributing samples and materials to this study. G.Smits and W.R. thank G. Kelsey and A. Ferguson-Smith for helpful comments, S. Andrews and I. McKendrick for bioinformatics help and F. Santos and M. Hinton for photographics help. This work was supported by the International Human Frontier Science Program Organisation (G.Smits), the Biotechnology and Biological Sciences Research Council and the Medical Research Council (W.R.), the European Union FP6 Network of Excellence “The Epigenome” (W.R., I.D.), the Wellcome Trust (A.J.M., S.G.-J., L.M., J.R., I.D.), the National Health and Medical Research Council (A.J.P) and the Australian Research Council (M.B.R.). The supporting funding agencies had no roles in the design and conduct of the study, in the collection, analysis, and interpretation of the data, and in the preparation, review, or approval of the manuscript.

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G.Smits., A.J.M., W.R. and I.D. planned the experiments and the writing of the manuscript; G.Smits, A.J.M., P.S. and D.B. performed the biochemical experiments; G.Smits, A.J.M. and S.G.-J. performed the bioinformatics analyses; A.J.M., L.M. and J.R. selected and sequenced the new BACs described in the text; A.J.P., G.Shaw and M.B.R. contributed the wallaby tissue samples; J.L.V. and J.R.M. contributed the opossum tissue samples. G.Smits and A.J.M. contributed equally to the study as first authors. W.R. and I.D. contributed equally to the study as senior authors. G.Smits and W.R. wrote the manuscript. All authors commented on the manuscript.

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Correspondence to Wolf Reik or Ian Dunham.

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Smits, G., Mungall, A., Griffiths-Jones, S. et al. Conservation of the H19 noncoding RNA and H19-IGF2 imprinting mechanism in therians. Nat Genet 40, 971–976 (2008). https://doi.org/10.1038/ng.168

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