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Loss of silent-chromatin looping and impaired imprinting of DLX5 in Rett syndrome

Abstract

Mutations in MECP2 are associated with Rett syndrome, an X-linked neurodevelopmental disorder. To identify genes targeted by Mecp2, we sequenced 100 in vivo Mecp2-binding sites in mouse brain. Several sequences mapped to an imprinted gene cluster on chromosome 6, including Dlx5 and Dlx6, whose transcription was roughly two times greater in brains of Mecp2-null mice compared with those of wild-type mice. The maternally expressed gene DLX5 showed a loss of imprinting in lymphoblastoid cells from individuals with Rett syndrome. Because Dlx5 regulates production of enzymes that synthesize γ-aminobutyric acid (GABA), loss of imprinting of Dlx5 may alter GABAergic neuron activity in individuals with Rett syndrome. In mouse brain, Dlx5 imprinting was relaxed, yet Mecp2-mediated silent-chromatin structure existed at the Dlx5-Dlx6 locus in brains of wild-type, but not Mecp2-null, mice. Mecp2 targeted histone deacetylase 1 to a sharply defined, 1-kb region at the Dlx5-Dlx6 locus and promoted repressive histone methylation at Lys9 at this site. Chromatin immunoprecipitation–combined loop assays showed that Mecp2 mediated the silent chromatin–derived 11-kb chromatin loop at the Dlx5-Dlx6 locus. This loop was absent in chromatin of brains of Mecp2-null mice, and Dlx5-Dlx6 interacted with far distant sequences, forming distinct active chromatin–associated loops. These results show that formation of a silent-chromatin loop is a new mechanism underlying gene regulation by Mecp2.

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Figure 1: Ablation of Mecp2 results in elevated transcription of Dlx5 and Dlx6 in the imprinted gene cluster of chromosome 6 containing in vivo MBSs.
Figure 2: Preferential transcription of Dlx5 from the maternal allele was lost in brains of Mecp2-null mice.
Figure 3: LOI of DLX5 in LCLs from individuals with RTT carrying mutations in MECP2.
Figure 4: Methylation patterns are similar in the Dlx5-Dlx6 locus in brains of wild-type and Mecp2-null mice.
Figure 5: Mecp2 recruits Hdac1 and mediates dimethylation at H3-Lys9 at the Dlx5-Dlx6 locus.
Figure 6: Mecp2 is required for organization of the silent-chromatin loop in the Dlx5-Dlx6 locus.

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Acknowledgements

We thank Y. Kohwi for valuable suggestions and F. Creegan, S. Krauss and the members of the laboratory for critical reading of the manuscript. This work was supported by an International Rett Syndrome Association Fellowship to S.H., a Rett Syndrome Research Foundation Fellowship to M.M. and by a US National Institutes of Health grant to T.K.-S.

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Correspondence to Terumi Kohwi-Shigematsu.

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Supplementary information

Supplementary Fig. 1

DLX5 expression levels and allelic expression of IGF2 and KCNQ1OT/LIT1 in LCLs from RTT patients. (PDF 468 kb)

Supplementary Fig. 2

Analysis of the methylation profiles of specific regions in the Dlx5/DLX5 locus. (PDF 821 kb)

Supplementary Table 1

Mecp2 binding sequences. (PDF 43 kb)

Supplementary Note

Classification of Mecp2-binding sequences. (PDF 46 kb)

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Horike, Si., Cai, S., Miyano, M. et al. Loss of silent-chromatin looping and impaired imprinting of DLX5 in Rett syndrome. Nat Genet 37, 31–40 (2005). https://doi.org/10.1038/ng1491

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