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In vivo analysis of Cajal body movement, separation, and joining in live human cells.

Platani M, Goldberg I, Swedlow JR, Lamond AI - J. Cell Biol. (2000)

Bottom Line: Cajal bodies are localized in the nucleoplasm and are often found at the nucleolar periphery.The GFP-coilin protein is dynamically associated with Cajal bodies as shown by changes in their fluorescence intensity over time.This study reveals an unexpectedly high level of movement and interactions of nuclear bodies in human cells and suggests that these movements may be driven, at least in part, by regulated mechanisms.

View Article: PubMed Central - PubMed

Affiliation: MSI/WTB Complex, University of Dundee, Dundee DD1 5EH, Scotland.

ABSTRACT
Cajal bodies (also known as coiled bodies) are subnuclear organelles that contain specific nuclear antigens, including splicing small nuclear ribonucleoproteins (snRNPs) and a subset of nucleolar proteins. Cajal bodies are localized in the nucleoplasm and are often found at the nucleolar periphery. We have constructed a stable HeLa cell line, HeLa(GFP-coilin), that expresses the Cajal body marker protein, p80 coilin, fused to the green fluorescent protein (GFP-coilin). The localization pattern and biochemical properties of the GFP-coilin fusion protein are identical to the endogenous p80 coilin. Time-lapse recordings on 63 nuclei of HeLa(GFP-coilin) cells showed that all Cajal bodies move within the nucleoplasm. Movements included translocations through the nucleoplasm, joining of bodies to form larger structures, and separation of smaller bodies from larger Cajal bodies. Also, we observed Cajal bodies moving to and from nucleoli. The data suggest that there may be at least two classes of Cajal bodies that differ in their size, antigen composition, and dynamic behavior. The smaller size class shows more frequent and faster rates of movement, up to 0.9 microm/min. The GFP-coilin protein is dynamically associated with Cajal bodies as shown by changes in their fluorescence intensity over time. This study reveals an unexpectedly high level of movement and interactions of nuclear bodies in human cells and suggests that these movements may be driven, at least in part, by regulated mechanisms.

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In vivo time-lapse three-dimensional analysis of an HeLaGFP-coilin cell nucleus showing movement of CBs through the nucleoplasm and separation of mini-CBs from Cajal bodies. 12 three-dimensional time-lapse images out of a total of 22 are shown as a montage. The image at each time point is a maximum intensity projection of 20 optical sections (0.5 μm each) scanning the entire cell nucleus. Data were collected every 3 min over a period of 2.25 h. Note the movement of a CB (magenta arrowheads) through the nucleoplasm from right to left bringing it into close proximity to a Cajal body that was located at the opposite side of the nucleus at time 0.00. A mini-CB (blue arrowhead) can be seen separating from a larger Cajal body located at the nuclear periphery, moving to the nucleolus and returning back to the same Cajal body from where it originated. An animated time-lapse version of this process is available at http://www.jcb.org/cgi/content/full/151/7/1561/DC1. Bar, 15 μm.
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Figure 6: In vivo time-lapse three-dimensional analysis of an HeLaGFP-coilin cell nucleus showing movement of CBs through the nucleoplasm and separation of mini-CBs from Cajal bodies. 12 three-dimensional time-lapse images out of a total of 22 are shown as a montage. The image at each time point is a maximum intensity projection of 20 optical sections (0.5 μm each) scanning the entire cell nucleus. Data were collected every 3 min over a period of 2.25 h. Note the movement of a CB (magenta arrowheads) through the nucleoplasm from right to left bringing it into close proximity to a Cajal body that was located at the opposite side of the nucleus at time 0.00. A mini-CB (blue arrowhead) can be seen separating from a larger Cajal body located at the nuclear periphery, moving to the nucleolus and returning back to the same Cajal body from where it originated. An animated time-lapse version of this process is available at http://www.jcb.org/cgi/content/full/151/7/1561/DC1. Bar, 15 μm.

Mentions: The shape of the nuclei shown in Fig. 2, Fig. 5, Fig. 6, and Fig. 7 are quite variable, even during the period of data collection. This nuclear morphology is a characteristic of both parental and HeLaGFP-coilin cell lines (see Fig. 2) and is not induced by phototoxicity or imaging. Cells with similar interphase morphology entered and completed mitosis (data not shown).


In vivo analysis of Cajal body movement, separation, and joining in live human cells.

Platani M, Goldberg I, Swedlow JR, Lamond AI - J. Cell Biol. (2000)

In vivo time-lapse three-dimensional analysis of an HeLaGFP-coilin cell nucleus showing movement of CBs through the nucleoplasm and separation of mini-CBs from Cajal bodies. 12 three-dimensional time-lapse images out of a total of 22 are shown as a montage. The image at each time point is a maximum intensity projection of 20 optical sections (0.5 μm each) scanning the entire cell nucleus. Data were collected every 3 min over a period of 2.25 h. Note the movement of a CB (magenta arrowheads) through the nucleoplasm from right to left bringing it into close proximity to a Cajal body that was located at the opposite side of the nucleus at time 0.00. A mini-CB (blue arrowhead) can be seen separating from a larger Cajal body located at the nuclear periphery, moving to the nucleolus and returning back to the same Cajal body from where it originated. An animated time-lapse version of this process is available at http://www.jcb.org/cgi/content/full/151/7/1561/DC1. Bar, 15 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2150679&req=5

Figure 6: In vivo time-lapse three-dimensional analysis of an HeLaGFP-coilin cell nucleus showing movement of CBs through the nucleoplasm and separation of mini-CBs from Cajal bodies. 12 three-dimensional time-lapse images out of a total of 22 are shown as a montage. The image at each time point is a maximum intensity projection of 20 optical sections (0.5 μm each) scanning the entire cell nucleus. Data were collected every 3 min over a period of 2.25 h. Note the movement of a CB (magenta arrowheads) through the nucleoplasm from right to left bringing it into close proximity to a Cajal body that was located at the opposite side of the nucleus at time 0.00. A mini-CB (blue arrowhead) can be seen separating from a larger Cajal body located at the nuclear periphery, moving to the nucleolus and returning back to the same Cajal body from where it originated. An animated time-lapse version of this process is available at http://www.jcb.org/cgi/content/full/151/7/1561/DC1. Bar, 15 μm.
Mentions: The shape of the nuclei shown in Fig. 2, Fig. 5, Fig. 6, and Fig. 7 are quite variable, even during the period of data collection. This nuclear morphology is a characteristic of both parental and HeLaGFP-coilin cell lines (see Fig. 2) and is not induced by phototoxicity or imaging. Cells with similar interphase morphology entered and completed mitosis (data not shown).

Bottom Line: Cajal bodies are localized in the nucleoplasm and are often found at the nucleolar periphery.The GFP-coilin protein is dynamically associated with Cajal bodies as shown by changes in their fluorescence intensity over time.This study reveals an unexpectedly high level of movement and interactions of nuclear bodies in human cells and suggests that these movements may be driven, at least in part, by regulated mechanisms.

View Article: PubMed Central - PubMed

Affiliation: MSI/WTB Complex, University of Dundee, Dundee DD1 5EH, Scotland.

ABSTRACT
Cajal bodies (also known as coiled bodies) are subnuclear organelles that contain specific nuclear antigens, including splicing small nuclear ribonucleoproteins (snRNPs) and a subset of nucleolar proteins. Cajal bodies are localized in the nucleoplasm and are often found at the nucleolar periphery. We have constructed a stable HeLa cell line, HeLa(GFP-coilin), that expresses the Cajal body marker protein, p80 coilin, fused to the green fluorescent protein (GFP-coilin). The localization pattern and biochemical properties of the GFP-coilin fusion protein are identical to the endogenous p80 coilin. Time-lapse recordings on 63 nuclei of HeLa(GFP-coilin) cells showed that all Cajal bodies move within the nucleoplasm. Movements included translocations through the nucleoplasm, joining of bodies to form larger structures, and separation of smaller bodies from larger Cajal bodies. Also, we observed Cajal bodies moving to and from nucleoli. The data suggest that there may be at least two classes of Cajal bodies that differ in their size, antigen composition, and dynamic behavior. The smaller size class shows more frequent and faster rates of movement, up to 0.9 microm/min. The GFP-coilin protein is dynamically associated with Cajal bodies as shown by changes in their fluorescence intensity over time. This study reveals an unexpectedly high level of movement and interactions of nuclear bodies in human cells and suggests that these movements may be driven, at least in part, by regulated mechanisms.

Show MeSH
Related in: MedlinePlus