The Cajal body (CB) is a conserved non-membrane nuclear structure where several steps of small nuclear RNP particle (snRNP) biogenesis take place. It has been proposed that CB formation follows a liquid-liquid phase separation model, but this hypothesis has never been rigorously tested. Here, we applied live-cell imaging to show that the key CB assembly factor coilin is mobile within the CB, and we revealed a diffusion barrier that limits the coilin exchange between CBs and the nucleoplasm. We generated single aa mutations and demonstrated that RNA-dependent coilin oligomerization and coilin interaction with snRNP are essential for CB formation and maintenance. We applied these data to formulate a mathematical model that links the movement of coilin within the nucleoplasm, CB, and across the boundary with its oligomerization and snRNP binding. Our results illustrate CB as a structure dynamically responding to snRNP assembly and recycling.

• MeSH
• Cajalova tělíska * metabolismus   genetika   MeSH
• HeLa buňky MeSH
• jaderné proteiny * metabolismus   genetika   MeSH
• lidé MeSH
• multimerizace proteinu MeSH
• mutace MeSH
• ribonukleoproteiny malé jaderné * metabolismus   genetika   MeSH
• spliceozomy * metabolismus   genetika   MeSH
• vazba proteinů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• jaderné proteiny * MeSH
• p80-coilin MeSH Prohlížeč
• ribonukleoproteiny malé jaderné * MeSH

The nucleus of higher eukaryotes contains a number of structures that concentrate specific biomolecules and play distinct roles in nuclear metabolism. In recent years, the molecular mechanisms controlling their formation have been intensively studied. In this brief review, I focus on coilin and Cajal bodies. Coilin is a key scaffolding protein of Cajal bodies that is evolutionarily conserved in metazoans. Cajal bodies are thought to be one of the archetypal nuclear structures involved in the metabolism of several short non-coding nuclear RNAs. Yet surprisingly little is known about the structure and function of coilin, and a comprehensive model to explain the origin of Cajal bodies is also lacking. Here, I summarize recent results on Cajal bodies and coilin and discuss them in the context of the last three decades of research in this field.

• Klíčová slova
• Cajal bodies, coilin, snRNA, snRNP, snoRNA, telomerase RNA,
• MeSH
• buněčné jádro * MeSH
• Cajalova tělíska * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

Nucleus, chromatin, and chromosome organization studies heavily rely on fluorescence microscopy imaging to elucidate the distribution and abundance of structural and regulatory components. Three-dimensional (3D) image stacks are a source of quantitative data on signal intensity level and distribution and on the type and shape of distribution patterns in space. Their analysis can lead to novel insights that are otherwise missed in qualitative-only analyses. Quantitative image analysis requires specific software and workflows for image rendering, processing, segmentation, setting measurement points and reference frames and exporting target data before further numerical processing and plotting. These tasks often call for the development of customized computational scripts and require an expertise that is not broadly available to the community of experimental biologists. Yet, the increasing accessibility of high- and super-resolution imaging methods fuels the demand for user-friendly image analysis workflows. Here, we provide a compendium of strategies developed by participants of a training school from the COST action INDEPTH to analyze the spatial distribution of nuclear and chromosomal signals from 3D image stacks, acquired by diffraction-limited confocal microscopy and super-resolution microscopy methods (SIM and STED). While the examples make use of one specific commercial software package, the workflows can easily be adapted to concurrent commercial and open-source software. The aim is to encourage biologists lacking custom-script-based expertise to venture into quantitative image analysis and to better exploit the discovery potential of their images.Abbreviations: 3D FISH: three-dimensional fluorescence in situ hybridization; 3D: three-dimensional; ASY1: ASYNAPTIC 1; CC: chromocenters; CO: Crossover; DAPI: 4',6-diamidino-2-phenylindole; DMC1: DNA MEIOTIC RECOMBINASE 1; DSB: Double-Strand Break; FISH: fluorescence in situ hybridization; GFP: GREEN FLUORESCENT PROTEIN; HEI10: HUMAN ENHANCER OF INVASION 10; NCO: Non-Crossover; NE: Nuclear Envelope; Oligo-FISH: oligonucleotide fluorescence in situ hybridization; RNPII: RNA Polymerase II; SC: Synaptonemal Complex; SIM: structured illumination microscopy; ZMM (ZIP: MSH4: MSH5 and MER3 proteins); ZYP1: ZIPPER-LIKE PROTEIN 1.

• Klíčová slova
• 3D organization, Nucleus, RNA Pol II, SIM, STED imaging, chromatin, chromosome, crossovers, image analysis, meiosis, metaphase, mitosis, nuclear bodies, nuclear speckles, oligo FISH, pachytene, quantification, segmentation, spatial distribution, transcription factories,
• MeSH
• buněčné jádro * MeSH
• chromatin * MeSH
• fluorescenční mikroskopie MeSH
• hybridizace in situ fluorescenční MeSH
• lidé MeSH
• průběh práce MeSH
• zelené fluorescenční proteiny MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chromatin * MeSH
• zelené fluorescenční proteiny MeSH

Understanding how the packaging of chromatin in the nucleus is regulated and organized to guide complex cellular and developmental programmes, as well as responses to environmental cues is a major question in biology. Technological advances have allowed remarkable progress within this field over the last years. However, we still know very little about how the 3D genome organization within the cell nucleus contributes to the regulation of gene expression. The nuclear space is compartmentalized in several domains such as the nucleolus, chromocentres, telomeres, protein bodies, and the nuclear periphery without the presence of a membrane around these domains. The role of these domains and their possible impact on nuclear activities is currently under intense investigation. In this review, we discuss new data from research in plants that clarify functional links between the organization of different nuclear domains and plant genome function with an emphasis on the potential of this organization for gene regulation.

• Klíčová slova
• 3D Chromatin organization, chromocentres, gene expression, liquid–liquid phase separation (LLPS), nuclear bodies, nuclear domains, nuclear periphery, nucleolus, telomeres, topologically associated domains (TADs),
• MeSH
• buněčné jadérko MeSH
• buněčné jádro * MeSH
• chromatin * MeSH
• regulace genové exprese MeSH
• rostliny genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• chromatin * MeSH