Abstract
Sugar beet (Beta vulgaris) chromosomes consist of large heterochromatic blocks in pericentromeric, centromeric, and intercalary regions comprised of two different highly abundant DNA satellite families. To investigate DNA methylation at single base resolution at heterochromatic regions, we applied a method for strand-specific bisulfite sequencing of more than 1,000 satellite monomers followed by statistical analyses. As a result, we uncovered diversity in the distribution of different methylation patterns in both satellite families. Heavily methylated CG and CHG (H=A, T, or C) sites occur more frequently in intercalary heterochromatin, while CHH sites, with the exception of CAA, are only sparsely methylated, in both intercalary and pericentromeric/centromeric heterochromatin. We show that the difference in DNA methylation intensity is correlated to unequal distribution of heterochromatic histone H3 methylation marks. While clusters of H3K9me2 were absent from pericentromeric heterochromatin and restricted only to intercalary heterochromatic regions, H3K9me1 and H3K27me1 were observed in all types of heterochromatin. By sequencing of a small RNA library consisting of 6.76 million small RNAs, we identified small interfering RNAs (siRNAs) of 24 nucleotides in size which originated from both strands of the satellite DNAs. We hypothesize an involvement of these siRNAs in the regulation of DNA and histone methylation for maintaining heterochromatin.
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Acknowledgments
Falk Zakrzewski acknowledges a fellowship and financial support of the FAZIT Foundation. This work is funded by the BMBF grant “Verbundprojekt GABI-Beet Physical map: Physikalische Genomkarte der Zuckerrübe zur Nutzung in der Pflanzenzüchtung”, FKZ 0313127B, 0313127D, and 0313127E (to B.W., H.H., T.S.); by the “Verbundprojekt GABI BeetSeq: Erstellung einer Referenzsequenz für das Genom der Zuckerrübe (Beta vulgaris)”, FKZ 0315069A and 0315069A (to H.H. and B.W.); and by a grant from the Land Sachsen-Anhalt W 21023908 (to J.F.). We thank Florian Mette for valuable discussion.
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ESM 1
(DOC 689 kb)
ESM 5
Data file (FASTA format) showing sequence alignment of satellite subfamily pBV I (RTF 106 kb)
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Data file (FASTA format) showing sequence alignment of satellite subfamily pBV II (RTF 255 kb)
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Data file (FASTA format) showing sequence alignment of satellite subfamily pBV III (RTF 553 kb)
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Data file (FASTA format) showing sequence alignment of satellite subfamily pBV IV (RTF 117 kb)
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Data file (FASTA format) showing sequence alignment of satellite subfamily pBV V (RTF 10 kb)
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Data file (FASTA format) showing sequence alignment of satellite subfamily pBV VI (RTF 13 kb)
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Data file (FASTA format) showing sequence alignment of satellite family pEV (RTF 4634 kb)
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Data file (FASTA format) showing sequence alignment of bisulfite-sequenced monomer regions of the forward strand of satellite subfamily pBV II (RTF 31 kb)
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Data file (FASTA format) showing sequence alignment of bisulfite-sequenced monomer regions of the complement strand of satellite subfamily pBV II (RTF 41 kb)
ESM 14
Data file (FASTA format) showing sequence alignment of bisulfite-sequenced monomer regions of the forward strand of satellite subfamily pBV III (RTF 150 kb)
ESM 15
Data file (FASTA format) showing sequence alignment of bisulfite-sequenced monomer regions of the complement strand of satellite subfamily pBV III (RTF 88 kb)
ESM 16
Data file (FASTA format) showing sequence alignment of bisulfite-sequenced monomer regions of the forward strand of satellite family pEV (RTF 139 kb)
ESM 17
Data file (FASTA format) showing sequence alignment of bisulfite-sequenced monomer regions of the complement strand of satellite family pEV (RTF 194 kb)
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Data file (FASTA format) showing sequence alignment of bisulfite-converted pGEM-T fragments (RTF 5 kb)
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Data file (FASTA format) showing small RNAs originating from satellites pBV and pEV (RTF 73 kb)
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Zakrzewski, F., Weisshaar, B., Fuchs, J. et al. Epigenetic profiling of heterochromatic satellite DNA. Chromosoma 120, 409–422 (2011). https://doi.org/10.1007/s00412-011-0325-x
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DOI: https://doi.org/10.1007/s00412-011-0325-x