title: The CHH motif in sugar beet satellite DNA: a modulator for cytosine methylation.
abstract: Methylation of DNA is important for the epigenetic silencing of repetitive DNA in plant genomes. Knowledge about the cytosine methylation status of satellite DNAs, a major class of repetitive DNA, is scarce. One reason for this is that arrays of tandemly arranged sequences are usually collapsed in next-generation sequencing assemblies. We applied strategies to overcome this limitation and quantified the level of cytosine methylation and its pattern in three satellite families of sugar beet (Beta vulgaris) which differ in their abundance, chromosomal localization and monomer size. We visualized methylation levels along pachytene chromosomes with respect to small satellite loci at maximum resolution using chromosome-wide fluorescent in situ hybridization complemented with immunostaining and super-resolution microscopy. Only reduced methylation of many satellite arrays was obtained. To investigate methylation at the nucleotide level we performed bisulfite sequencing of 1569 satellite sequences. We found that the level of methylation of cytosine strongly depends on the sequence context: cytosines in the CHH motif show lower methylation (44-52%), while CG and CHG motifs are more strongly methylated. This affects the overall methylation of satellite sequences because CHH occurs frequently while CG and CHG are rare or even absent in the satellite arrays investigated. Evidently, CHH is the major target for modulation of the cytosine methylation level of adjacent monomers within individual arrays and contributes to their epigenetic function. This strongly indicates that asymmetric cytosine methylation plays a role in the epigenetic modification of satellite repeats in plant genomes.
authors.affiliation: Department of Plant Cell and Molecular Biology, TU Dresden, D-01062, Dresden, Germany.
authors.forename: Falk
authors.initials: F
authors.lastname: Zakrzewski
authors.forename: Veit
authors.initials: V
authors.lastname: Schubert
authors.forename: Prisca
authors.initials: P
authors.lastname: Viehoever
authors.forename: André E
authors.initials: AE
authors.lastname: Minoche
authors.forename: Juliane C
authors.initials: JC
authors.lastname: Dohm
authors.forename: Heinz
authors.initials: H
authors.lastname: Himmelbauer
authors.forename: Bernd
authors.initials: B
authors.lastname: Weisshaar
authors.forename: Thomas
authors.initials: T
authors.lastname: Schmidt
mesh_terms: D027461:Beta vulgaris / Q000235:genetics*; D032461:Chromosomes, Plant; D003596:Cytosine / Q000378:metabolism*; D019175:DNA Methylation; D018744:DNA, Plant / Q000737:chemistry*; D044127:Epigenesis, Genetic; D018745:Genome, Plant; D059372:Nucleotide Motifs; D012091:Repetitive Sequences, Nucleic Acid; D017422:Sequence Analysis, DNA
chemical_list: D018744:DNA, Plant; D003596:Cytosine
keywords: Beta vulgaris; DNA methylation; bisulfite sequencing; satellite DNA; siRNAs; sugar beet; super-resolution microscopy