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Mammalian chromosomes are organized in structural and functional domains of 0.1–10 Mb, which are characterized by high self-association frequencies in the nuclear space and different contact probabilities with nuclear sub-compartments. They exhibit distinct chromatin modification patterns, gene expression levels and replication timing. Recently, nucleolus-associated chromosomal domains (NADs) have been discovered, yet their precise genomic organization and dynamics are still largely unknown. Here, we use nucleolus genomics and single-cell experiments to address these questions in human embryonic fibroblasts during replicative senescence. Genome-wide mapping reveals 1,646 NADs in proliferating cells, which cover about 38% of the annotated human genome. They are mainly heterochromatic and correlate with late replicating loci. Using Hi-C data analysis, we show that interactions of NADs dominate interphase chromosome contacts in the 10–50 Mb distance range. Interestingly, only minute changes in nucleolar association are observed upon senescence. These spatial rearrangements in subdomains smaller than 100 kb are accompanied with local transcriptional changes. In contrast, large centromeric and pericentromeric satellite repeat clusters extensively dissociate from nucleoli in senescent cells. Accordingly, H3K9me3-marked heterochromatin gets remodelled at the perinucleolar space as revealed by immunofluorescence analyses. Collectively, this study identifies connections between the nucleolus, 3D genome structure, and cellular aging at the level of interphase chromosome organization. [ABSTRACT FROM AUTHOR]

Upstream Binding Factor (UBF) is a unique multi-HMGB-box protein first identified as a co-factor in RNA polymerase I (RPI/PolI) transcription. However, its poor DNA sequence selectivity and its ability to generate nucleosome-like nucleoprotein complexes suggest a more generalized role in chromatin structure. We previously showed that extensive depletion of UBF reduced the number of actively transcribed ribosomal RNA (rRNA) genes, but had little effect on rRNA synthesis rates or cell proliferation, leaving open the question of its requirement for RPI transcription. Using gene deletion in mouse, we now show that UBF is essential for embryo development beyond morula. Conditional deletion in cell cultures reveals that UBF is also essential for transcription of the rRNA genes and that it defines the active chromatin conformation of both gene and enhancer sequences. Loss of UBF prevents formation of the SL1/TIF1B pre-initiation complex and recruitment of the RPI-Rrn3/TIF1A complex. It is also accompanied by recruitment of H3K9me3, canonical histone H1 and HP1α, but not by de novo DNA methylation. Further, genes retain penta-acetyl H4 and H2A.Z, suggesting that even in the absence of UBF the rRNA genes can maintain a potentially active state. In contrast to canonical histone H1, binding of H1.4 is dependent on UBF, strongly suggesting that it plays a positive role in gene activity. Unexpectedly, arrest of rRNA synthesis does not suppress transcription of the 5S, tRNA or snRNA genes, nor expression of the several hundred mRNA genes implicated in ribosome biogenesis. Thus, rRNA gene activity does not coordinate global gene expression for ribosome biogenesis. Loss of UBF also unexpectedly induced the formation in cells of a large sub-nuclear structure resembling the nucleolar precursor body (NPB) of oocytes and early embryos. These somatic NPBs contain rRNA synthesis and processing factors but do not associate with the rRNA gene loci (NORs). [ABSTRACT FROM AUTHOR]