Karina Zillner1, Jun Komatsu2, Katharina Filarsky1,3, Rajakiran Kalepu1,4, Aaron Bensimon2, Attila Németh1. 1. Department of Biochemistry III, Biochemistry Center Regensburg, University of Regensburg, Universitätsstraße 31, 93053 Regensburg, Germany. 2. Genomic Vision, 80 Rue des Meuniers, 92220 Bagneux, France. 3. German Cancer Research Center (DKFZ), Heidelberg 69120, Germany. 4. University Hospital Ulm, Ulm 89070, Germany.
Abstract
AIM: The synthesis of rRNA is a key determinant of normal and malignant cell growth and subject to epigenetic regulation. Yet, the epigenomic features of rDNA arrays clustered in nucleolar organizer regions are largely unknown. We set out to explore for the first time how DNA methylation is distributed on individual rDNA arrays. MATERIALS & METHODS: Here we combined immunofluorescence detection of DNA modifications with fluorescence hybridization of single DNA fibers, metaphase immuno-FISH and methylation-sensitive restriction enzyme digestions followed by Southern blot. RESULTS: We found clustering of both hypomethylated and hypermethylated repeat units and hypermethylation of noncanonical rDNA in IMR90 fibroblasts and HCT116 colorectal carcinoma cells. Surprisingly, we also found transitions between hypo- and hypermethylated rDNA repeat clusters on single DNA fibers. CONCLUSION: Collectively, our analyses revealed co-existence of different epialleles on individual nucleolar organizer regions and showed that epi-combing is a valuable approach to analyze epigenomic patterns of repetitive DNA.
AIM: The synthesis of rRNA is a key determinant of normal and malignant cell growth and subject to epigenetic regulation. Yet, the epigenomic features of rDNA arrays clustered in nucleolar organizer regions are largely unknown. We set out to explore for the first time how DNA methylation is distributed on individual rDNA arrays. MATERIALS & METHODS: Here we combined immunofluorescence detection of DNA modifications with fluorescence hybridization of single DNA fibers, metaphase immuno-FISH and methylation-sensitive restriction enzyme digestions followed by Southern blot. RESULTS: We found clustering of both hypomethylated and hypermethylated repeat units and hypermethylation of noncanonical rDNA in IMR90 fibroblasts and HCT116 colorectal carcinoma cells. Surprisingly, we also found transitions between hypo- and hypermethylated rDNA repeat clusters on single DNA fibers. CONCLUSION: Collectively, our analyses revealed co-existence of different epialleles on individual nucleolar organizer regions and showed that epi-combing is a valuable approach to analyze epigenomic patterns of repetitive DNA.
Authors: Bo Li; Karl A G Kremling; Penghao Wu; Robert Bukowski; Maria C Romay; En Xie; Edward S Buckler; Mingsheng Chen Journal: Genome Res Date: 2018-08-30 Impact factor: 9.043
Authors: Sam Kint; Wim Van Criekinge; Linos Vandekerckhove; Winnok H De Vos; Karol Bomsztyk; Diane S Krause; Oleg Denisenko Journal: Nucleic Acids Res Date: 2021-05-07 Impact factor: 16.971