Identifying gene locus associations with promyelocytic leukemia nuclear bodies using immuno-TRAP

Ching, Reagan W.; Ahmed, Kashif; Boutros, Paul C.; Penn, Linda Z.; Bazett-Jones, David P.

doi:10.1083/jcb.201211097

Cited by 43 publications

(64 citation statements)

References 34 publications

(41 reference statements)

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“…After formation at highly expressed target gene loci, CBs, and NBs in general, appear to attract other target genes that have localized to the interchromatin space and stably retain these regions in their periphery [30, 37, 39, 40, 60]. This ultimately influences global chromatin topology by forming nonrandom long-range gene pairing events.…”

Section: Gene Clusters Near Cajal Bodies Are Rna-dependentmentioning

confidence: 99%

“…Additionally, several NBs frequently form at specific genomic loci and accelerate target molecular processes in a localized microenvironment [1]. For example, nucleoli are formed around actively transcribing, tandemly-repeated ribosomal genes concentrated on several separate chromosomes [37], HLBs form at the replication-dependent histone gene clusters [38] and PML NBs also associate with defined genomic regions (albeit in a cell type-specific manner) [39]. However, NBs are multifunctional structures that influence the DNA damage response, protein and RNA modifications, as well as gene activity [1].…”

Section: Introductionmentioning

confidence: 99%

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Cajal body function in genome organization and transcriptome diversity

et al. 2016

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Nuclear bodies contribute to nonrandom organization of the human genome and nuclear function. Using a major prototypical nuclear body, the Cajal body, as an example, we suggest that these structures assemble at specific gene loci located across the genome as a result of high transcriptional activity. Subsequently, target genes are physically clustered in close proximity in Cajal body-containing cells. However, Cajal bodies are observed in only a limited number of human cell types, including neuronal and cancer cells. Ultimately, Cajal body depletion perturbs splicing kinetics by reducing target small nuclear RNA (snRNA) transcription and limiting the levels of spliceosomal snRNPs, including their modification and turnover following each round of RNA splicing. As such, Cajal bodies are capable of shaping the chromatin interaction landscape and the transcriptome by influencing spliceosome kinetics. Future studies should concentrate on characterizing the direct influence of Cajal bodies upon small nuclear RNA gene transcriptional dynamics.

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Section: Gene Clusters Near Cajal Bodies Are Rna-dependentmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Cajal body function in genome organization and transcriptome diversity

et al. 2016

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“…One of these sub-compartments is the nuclear lamina; the lamina-associated domains (LADs) are large genomic regions that are generally transcriptionally repressed and are located along the nuclear envelope (Guelen et al 2008). Other nuclear structures that associate with specific loci include the nucleolus (Nemeth and Langst 2011;Olson and Dundr 2001), transcription factories (Osborne et al 2004) and a variety of nuclear bodies, such as promyelocytic leukaemia (PML) nuclear bodies (Ching et al 2013), Cajal bodies (Smith and Lawrence 2000) and histone locus bodies (Liu et al 2006;Nizami et al 2010). This spatial proximity between specific loci and nuclear factors or sub-compartments may be a consequence of either active movement directed by nuclear skeletal elements (Bridger 2011;Bridger et al 2014;Chuang et al 2006;Dundr et al 2007) or may reflect a passive mass action and selfassembly of functional domains that stabilize once formed (reviewed in Chuang and Belmont 2007).…”

Section: Higher Order Chromatin Structure and Organizationmentioning

confidence: 99%

Coming to terms with chromatin structure

et al. 2015

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Chromatin, once thought to serve only as a means to package DNA, is now recognized as a major regulator of gene activity. As a result of the wide range of methods used to describe the numerous levels of chromatin organization, the terminology that has emerged to describe these organizational states is often imprecise and sometimes misleading. In this review, we discuss our current understanding of chromatin architecture and propose terms to describe the various biochemical and structural states of chromatin.

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“…Intriguingly, several sequencing-based studies have recently indicated that NBs assemble as a direct consequence of specific gene expression activity and form characteristic long-lived contact interactions with a defined cohort of genomic loci and CTs. [19][20][21] Nonetheless, the limitations of these populationbased studies using millions of mostly asynchronous cells are becoming apparent. Despite indications that genomic domains are known to form and maintain numerous spatial associations with both near (kBp) and distant (MBp) genomic locations, 22 it is unknown how many physical gene pairing events occur simultaneously.…”

Section: Introductionmentioning

confidence: 99%

Spectral imaging to visualize higher-order genomic organization

Sawyer

Shevtsov

Dundr

2016

Nucleus

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A concern in the field of genomics is the proper interpretation of large, high-throughput sequencing datasets. The use of DNA FISH followed by high-content microscopy is a valuable tool for validation and contextualization of frequently occurring gene pairing events at the single-cell level identified by deep sequencing. However, these techniques possess certain limitations. Firstly, they do not permit the study of colocalization of many gene loci simultaneously. Secondly, the direct assessment of the relative position of many clustered gene loci within their respective chromosome territories is impossible. Thus, methods are required to advance the study of higher-order nuclear and cellular organization. Here, we describe a multiplexed DNA FISH technique combined with indirect immunofluorescence to study the relative position of 6 distinct genomic or cellular structures. This can be achieved in a single hybridization step using spectral imaging during image acquisition and linear unmixing. Here, we detail the use of this method to quantify gene pairing between highly expressed spliceosomal genes and compare these data to randomly positioned in silico simulated gene clusters. This is a potentially universally applicable approach for the validation of 3C-based technologies, deep imaging of spatial organization within the nucleus and global cellular organization.

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Identifying gene locus associations with promyelocytic leukemia nuclear bodies using immuno-TRAP

Abstract: A new immuno-TRAP technique overcomes limitations of spatial resolution and selection bias to identify gene locus associations with a nuclear subcompartment such as promyelocytic leukemia nuclear bodies.

Cited by 43 publications

References 34 publications

Cajal body function in genome organization and transcriptome diversity

Cajal body function in genome organization and transcriptome diversity

Coming to terms with chromatin structure

Spectral imaging to visualize higher-order genomic organization

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