Reconstitution of human rRNA gene transcription in mouse cells by complete SL1 complex

Murano, Kensaku; Okuwaki, Mitsuru; Momose, Fumitaka; Kumakura, Michiko; Ueshima, Shuhei; Newbold, Robert F.; Nagata, Kyosuke

doi:10.1242/jcs.146787

Cited by 11 publications

(14 citation statements)

References 55 publications

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“…However, whether these contact sites would correspond to two identical SL1 complexes, or to two SL1 subcomplexes as seen in yeast, where distinct TAF1 subcomplexes bind UPE and core elements and are bridged by TBP ( Moss et al 2007 ), will require further study. It is relevant here to note that our present knowledge of the structure of mammalian SL1 is still incomplete ( Gorski et al 2007 ; Murano et al 2014 ).…”

Section: Discussionmentioning

confidence: 99%

A Deconvolution Protocol for ChIP-Seq Reveals Analogous Enhancer Structures on the Mouse and Human Ribosomal RNA Genes

Mars

Sabourin-Felix

Tremblay³

et al. 2018

G3 Genes|Genomes|Genetics

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The combination of Chromatin Immunoprecipitation and Massively Parallel Sequencing, or ChIP-Seq, has greatly advanced our genome-wide understanding of chromatin and enhancer structures. However, its resolution at any given genetic locus is limited by several factors. In applying ChIP-Seq to the study of the ribosomal RNA genes, we found that a major limitation to resolution was imposed by the underlying variability in sequence coverage that very often dominates the protein–DNA interaction profiles. Here, we describe a simple numerical deconvolution approach that, in large part, corrects for this variability, and significantly improves both the resolution and quantitation of protein–DNA interaction maps deduced from ChIP-Seq data. This approach has allowed us to determine the in vivo organization of the RNA polymerase I preinitiation complexes that form at the promoters and enhancers of the mouse (Mus musculus) and human (Homo sapiens) ribosomal RNA genes, and to reveal a phased binding of the HMG-box factor UBF across the rDNA. The data identify and map a “Spacer Promoter” and associated stalled polymerase in the intergenic spacer of the human ribosomal RNA genes, and reveal a very similar enhancer structure to that found in rodents and lower vertebrates.

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Section: Discussionmentioning

confidence: 99%

A Deconvolution Protocol for ChIP-Seq Reveals Analogous Enhancer Structures on the Mouse and Human Ribosomal RNA Genes

Mars

Sabourin-Felix

Tremblay³

et al. 2018

G3 Genes|Genomes|Genetics

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“…For example, mouse SL1 cannot form PICs on human promoters either in vitro or in the context of mouse monochromosomal somatic cell hybrids Sullivan et al 2001). Introduction of human TAF I s into mouse somatic cell hybrids can reactivate human NORs (Murano et al 2014). Nucleolar dominance between more closely related species is more difficult to explain.…”

Section: Nors As Units Of Regulation-a Role For Chromosomal Contextmentioning

confidence: 99%

Nucleolar organizer regions: genomic ‘dark matter’ requiring illumination

2016

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Nucleoli form around tandem arrays of a ribosomal gene repeat, termed nucleolar organizer regions (NORs). During metaphase, active NORs adopt a characteristic undercondensed morphology. Recent evidence indicates that the HMG-box-containing DNA-binding protein UBF (upstream binding factor) is directly responsible for this morphology and provides a mitotic bookmark to ensure rapid nucleolar formation beginning in telophase in human cells. This is likely to be a widely employed strategy, as UBF is present throughout metazoans. In higher eukaryotes, NORs are typically located within regions of chromosomes that form perinucleolar heterochromatin during interphase. Typically, the genomic architecture of NORs and the chromosomal regions within which they lie is very poorly described, yet recent evidence points to a role for context in their function. In Arabidopsis, NOR silencing appears to be controlled by sequences outside the rDNA (ribosomal DNA) array. Translocations reveal a role for context in the expression of the NOR on the X chromosome in Drosophila. Recent work has begun on characterizing the genomic architecture of human NORs. A role for distal sequences located in perinucleolar heterochromatin has been inferred, as they exhibit a complex transcriptionally active chromatin structure. Links between rDNA genomic stability and aging in Saccharomyces cerevisiae are now well established, and indications are emerging that this is important in aging and replicative senescence in higher eukaryotes. This, combined with the fact that rDNA arrays are recombinational hot spots in cancer cells, has focused attention on DNA damage responses in NORs. The introduction of DNA double-strand breaks into rDNA arrays leads to a dramatic reorganization of nucleolar structure. Damaged rDNA repeats move from the nucleolar interior to form caps at the nucleolar periphery, presumably to facilitate repair, suggesting that the chromosomal context of human NORs contributes to their genomic stability. The inclusion of NORs and their surrounding chromosomal environments in future genome drafts now becomes a priority.The relationship between nucleolar organizer regions (NORs) and nucleoli was first established in the 1930s (Heitz 1931;McClintock 1934), but, for decades, the content of the former and the role of the latter remained mysterious. The era of molecular and cellular biology revealed that NORs contain tandem arrays of ribosomal gene (rDNA) repeats and that nucleoli are the sites of ribosome biogenesis. Biochemistry has revealed the inner workings of the nucleolus and the complexity of ribosome biogenesis (for review, see Pederson 2010). However, the genomic architecture of NORs and the chromosomal context in which they lie remains undetermined for most eukaryotes. The resulting void has placed limitations on our understanding of the fundamental mechanisms by which NORs orchestrate formation of the largest structure in the nucleus. In this review, I discuss recent findings regarding the morphology of active NORs and how they seed r...

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“…The basal factors of the mammalian RPI transcription machinery include Selectivity Factor 1 (SL1), the multi-HMGbox factor UBTF/UBF and the RPI-associated initiation factor RRN3 [9,15,16] (Fig 1A). SL1 consists of the TATA-box Binding Protein (TBP) and the RPI specific TBP Associated Factors, TAFIA to D. Evolutionary variability of these TAFs determine their species-specific functions and that of RPI transcription, and for example human or mouse SL1 complexes are not functionally exchangeable [17][18][19]. In contrast, UBTF is a highly conserved essential factor that is functionally exchangeable between human, mouse and to some extent even Xenopus [19][20][21].…”

Section: Introductionmentioning

confidence: 99%

Ribosomal DNA promoter recognition is determined in vivo by cooperation between UBTF1 and SL1 and is compromised in the UBTF-E210K neuroregression syndrome

Tremblay¹,

Sibai

Valere

et al. 2022

PLoS Genet

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Transcription of the ~200 mouse and human ribosomal RNA genes (rDNA) by RNA Polymerase I (RPI/PolR1) accounts for 80% of total cellular RNA, around 35% of all nuclear RNA synthesis, and determines the cytoplasmic ribosome complement. It is therefore a major factor controlling cell growth and its misfunction has been implicated in hypertrophic and developmental disorders. Activation of each rDNA repeat requires nucleosome replacement by the architectural multi-HMGbox factor UBTF to create a 15.7 kbp nucleosome free region (NFR). Formation of this NFR is also essential for recruitment of the TBP-TAFI factor SL1 and for preinitiation complex (PIC) formation at the gene and enhancer-associated promoters of the rDNA. However, these promoters show little sequence commonality and neither UBTF nor SL1 display significant DNA sequence binding specificity, making what drives PIC formation a mystery. Here we show that cooperation between SL1 and the longer UBTF1 splice variant generates the specificity required for rDNA promoter recognition in cell. We find that conditional deletion of the TAF1B subunit of SL1 causes a striking depletion of UBTF at both rDNA promoters but not elsewhere across the rDNA. We also find that while both UBTF1 and -2 variants bind throughout the rDNA NFR, only UBTF1 is present with SL1 at the promoters. The data strongly suggest an induced-fit model of RPI promoter recognition in which UBTF1 plays an architectural role. Interestingly, a recurrent UBTF-E210K mutation and the cause of a pediatric neurodegeneration syndrome provides indirect support for this model. E210K knock-in cells show enhanced levels of the UBTF1 splice variant and a concomitant increase in active rDNA copies. In contrast, they also display reduced rDNA transcription and promoter recruitment of SL1. We suggest the underlying cause of the UBTF-E210K syndrome is therefore a reduction in cooperative UBTF1-SL1 promoter recruitment that may be partially compensated by enhanced rDNA activation.

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Reconstitution of human rRNA gene transcription in mouse cells by complete SL1 complex

Cited by 11 publications

References 55 publications

A Deconvolution Protocol for ChIP-Seq Reveals Analogous Enhancer Structures on the Mouse and Human Ribosomal RNA Genes

A Deconvolution Protocol for ChIP-Seq Reveals Analogous Enhancer Structures on the Mouse and Human Ribosomal RNA Genes

Nucleolar organizer regions: genomic ‘dark matter’ requiring illumination

Ribosomal DNA promoter recognition is determined in vivo by cooperation between UBTF1 and SL1 and is compromised in the UBTF-E210K neuroregression syndrome

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