Characterisation of transcriptionally active and inactive chromatin domains in neurons

Abstract

The tandemly organised ribosomal DNA (rDNA) repeats are transcribed by a dedicated RNA polymerase in a specialised nuclear compartment, the nucleolus. There appears to be an intimate link between the maintenance of nucleolar structure and the presence of heterochromatic chromatin domains. This is particularly evident in many large neurons, where a single nucleolus is present, which is separated from the remainder of the nucleus by a characteristic shell of heterochromatin. Using a combined fluorescence in situ hybridisation and immunocytochemistry approach, we have analysed the molecular composition of this highly organised neuronal chromatin, to investigate its functional significance. We find that clusters of inactive, methylated rDNA repeats are present inside large neuronal nucleoli, which are often attached to the shell of heterochromatic DNA. Surprisingly, the methylated DNA-binding protein MeCP2, which is abundantly present in the centromeric and perinucleolar heterochromatin, does not associate significantly with the methylated rDNA repeats, whereas histone H1 does overlap partially with these clusters. Histone H1 also defines other, centromere-associated chromatin subdomains, together with the mammalian Polycomb group factor Eed. These data indicate that neuronal, perinucleolar heterochromatin consists of several classes of inactive DNA, that are linked to a fraction of the inactive rDNA repeats. These distinct chromatin domains may serve to regulate RNA transcription and processing efficiently and to protect rDNA repeats against unwanted silencing and/or homologous recombination events.

DNA Binding of Methyl-CpG-Binding Protein MeCP2 in Human MCF7 Cells

“…Immunostaining of MeCP2 again exhibited a granular pattern thoughout the nucleus, except for the nucleoli (Figure 5A). Sparing of the nucleoli from MeCP2 distribution, which is consistent with previous observations (8), was also verified by double immunostaining with MeCP2 and nucleolin antibodies (data not shown). Furthermore, MeCP2 did not preferentially localize to DAPI-stained shells of heterochromatin.…”

“…In murine metaphase chromosomes, MeCP2 is enriched in pericentromeric heterochromatin, in accordance with its content of major satellite DNA, the largest fraction of methylated sequences in mice (2). In interphase nuclei of murine neuronal cells, MeCP2 is associated with pericentromeric and perinucleolar heterochromatin but does not bind significantly to the heavily methylated ribosomal DNA repeats (8). The occupancy of specific genes by MeCP2 has been analyzed using chromatin immunoprecipitation assays.…”

The Role of rDNA Clusters in Global Epigenetic Gene Regulation

“…The number of FC-DFC modules is relatively constant for a particular cell type but differs widely between cell types, making it a powerful biomarker for cell classification (Lafontaine et al, 2021). Some rDNA clusters are silent and form constitutive heterochromatin and are not associated with nucleoli (Akhmanova et al, 2000). Active rDNA units, which represent about half of the rDNA copies, are bound with upstream binding factor (UBF) and form nucleoli.…”

Phospho-Tau and Chromatin Landscapes in Early and Late Alzheimer’s Disease

“…These sequences can reposition chromatin regions close to the nuclear envelope to silence genes [69]. In addition, a structural and functional relationship between tau and AT-rich alpha satellite repetitive sequences is involved in the organization of the centromeric heterochromatin ring from the nucleolusassociated domains (NADs) [53,[70][71][72]. This partly explains the early observation of tau in the nucleolus.…”