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The transcriptional repressor Kaiso localizes at the mitotic spindle and is a constituent of the pericentriolar material.

Soubry A, Staes K, Parthoens E, Noppen S, Stove C, Bogaert P, van Hengel J, van Roy F - PLoS ONE (2010)

Bottom Line: In the present study we monitored Kaiso's subcellular localization during the cell cycle and found the following: (1) during interphase, Kaiso is located not only in the nucleus, but also on microtubular structures, including the centrosome; (2) at metaphase, it is present at the centrosomes and on the spindle microtubules; (3) during telophase, it accumulates at the midbody.We found that Kaiso is a genuine PCM component that belongs to a pericentrin molecular complex.Knockdown of Kaiso accelerated cell proliferation.

View Article: PubMed Central - PubMed

Affiliation: Department for Molecular Biomedical Research, VIB, Ghent, Belgium.

ABSTRACT
Kaiso is a BTB/POZ zinc finger protein known as a transcriptional repressor. It was originally identified through its in vitro association with the Armadillo protein p120ctn. Subcellular localization of Kaiso in cell lines and in normal and cancerous human tissues revealed that its expression is not restricted to the nucleus. In the present study we monitored Kaiso's subcellular localization during the cell cycle and found the following: (1) during interphase, Kaiso is located not only in the nucleus, but also on microtubular structures, including the centrosome; (2) at metaphase, it is present at the centrosomes and on the spindle microtubules; (3) during telophase, it accumulates at the midbody. We found that Kaiso is a genuine PCM component that belongs to a pericentrin molecular complex. We analyzed the functions of different domains of Kaiso by visualizing the subcellular distribution of GFP-tagged Kaiso fragments throughout the cell cycle. Our results indicate that two domains are responsible for targeting Kaiso to the centrosomes and microtubules. The first domain, designated SA1 for spindle-associated domain 1, is located in the center of the Kaiso protein and localizes at the spindle microtubules and centrosomes; the second domain, SA2, is an evolutionarily conserved domain situated just before the zinc finger domain and might be responsible for localizing Kaiso towards the centrosomal region. Constructs containing both SA domains and Kaiso's aminoterminal BTB/POZ domain triggered the formation of abnormal centrosomes. We also observed that overexpression of longer or full-length Kaiso constructs led to mitotic cell arrest and frequent cell death. Knockdown of Kaiso accelerated cell proliferation. Our data reveal a new target for Kaiso at the centrosomes and spindle microtubules during mitosis. They also strongly imply that Kaiso's function as a transcriptional regulator might be linked to the control of the cell cycle and to cell proliferation in cancer.

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The GFP-tagged Kaiso fragments K3 and K7 abnormally localize at the centrosomes and cause aberrant chromosomal distribution.HEK293 cells were transfected with constructs GFP-K3 and GFP-K7 (see also Fig. 1). In the upper row, cells were imaged by focusing on, respectively, the upper centrosomal region (left) and the lower centrosomal region (middle) (see also insets for magnification of these regions); an overlay of the upper middle picture with DAPI staining to detect DNA is shown on the right. Pictures in the lower two rows also reveal abnormal chromosomal distributions upon overexpression of, respectively, Kaiso fragments K3 and K7. DNA was stained with DAPI. A Zeiss Axiophot microscope was used (100× objective lens).
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pone-0009203-g009: The GFP-tagged Kaiso fragments K3 and K7 abnormally localize at the centrosomes and cause aberrant chromosomal distribution.HEK293 cells were transfected with constructs GFP-K3 and GFP-K7 (see also Fig. 1). In the upper row, cells were imaged by focusing on, respectively, the upper centrosomal region (left) and the lower centrosomal region (middle) (see also insets for magnification of these regions); an overlay of the upper middle picture with DAPI staining to detect DNA is shown on the right. Pictures in the lower two rows also reveal abnormal chromosomal distributions upon overexpression of, respectively, Kaiso fragments K3 and K7. DNA was stained with DAPI. A Zeiss Axiophot microscope was used (100× objective lens).

Mentions: Time-lapse microscopy was used to follow HEK293 cells transfected with full-length GFP-tagged Kaiso (construct K5) after cell cycle synchronization. Premitotic cell cycle arrest was observed in 95% of the cells. Indeed, mitotic cells were rarely observed (exemplified in Fig. 6), as cells generally started to round up and died before mitosis could start. Time-lapse recording of this process is available online (http://www.dmbr.ugent.be/ext/public/publications/Soubry_2009/GFP_FL-Kaiso.avi) as a Suppl. Movie S1. Sixty hours after transfection, all the cells transfected with the full-length Kaiso were dead. In contrast, transfection with shorter constructs containing the SA1 or SA2 domain but not the complete BTB/POZ or zinc finger domain, such as K4, K8 and K9 (Fig. 1), caused a loss of no more than 10% of the cells. Constructs expressing a complete or nearly complete POZ/BTB, such as K1, K2, K3 and K7 (Fig. 1), caused death of 60 to 95% of the cells. K3 and K7 fragments contain both SA domains, and the surviving fraction of cells overexpressing these fragments showed during mitosis multiple, strong and aberrant dots at putative centrosomal regions (Fig. 9). Aberrant distribution of the chromosomes (Fig. 9, DAPI stain) was accompanied with these abnormally shaped structures.


The transcriptional repressor Kaiso localizes at the mitotic spindle and is a constituent of the pericentriolar material.

Soubry A, Staes K, Parthoens E, Noppen S, Stove C, Bogaert P, van Hengel J, van Roy F - PLoS ONE (2010)

The GFP-tagged Kaiso fragments K3 and K7 abnormally localize at the centrosomes and cause aberrant chromosomal distribution.HEK293 cells were transfected with constructs GFP-K3 and GFP-K7 (see also Fig. 1). In the upper row, cells were imaged by focusing on, respectively, the upper centrosomal region (left) and the lower centrosomal region (middle) (see also insets for magnification of these regions); an overlay of the upper middle picture with DAPI staining to detect DNA is shown on the right. Pictures in the lower two rows also reveal abnormal chromosomal distributions upon overexpression of, respectively, Kaiso fragments K3 and K7. DNA was stained with DAPI. A Zeiss Axiophot microscope was used (100× objective lens).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2821401&req=5

pone-0009203-g009: The GFP-tagged Kaiso fragments K3 and K7 abnormally localize at the centrosomes and cause aberrant chromosomal distribution.HEK293 cells were transfected with constructs GFP-K3 and GFP-K7 (see also Fig. 1). In the upper row, cells were imaged by focusing on, respectively, the upper centrosomal region (left) and the lower centrosomal region (middle) (see also insets for magnification of these regions); an overlay of the upper middle picture with DAPI staining to detect DNA is shown on the right. Pictures in the lower two rows also reveal abnormal chromosomal distributions upon overexpression of, respectively, Kaiso fragments K3 and K7. DNA was stained with DAPI. A Zeiss Axiophot microscope was used (100× objective lens).
Mentions: Time-lapse microscopy was used to follow HEK293 cells transfected with full-length GFP-tagged Kaiso (construct K5) after cell cycle synchronization. Premitotic cell cycle arrest was observed in 95% of the cells. Indeed, mitotic cells were rarely observed (exemplified in Fig. 6), as cells generally started to round up and died before mitosis could start. Time-lapse recording of this process is available online (http://www.dmbr.ugent.be/ext/public/publications/Soubry_2009/GFP_FL-Kaiso.avi) as a Suppl. Movie S1. Sixty hours after transfection, all the cells transfected with the full-length Kaiso were dead. In contrast, transfection with shorter constructs containing the SA1 or SA2 domain but not the complete BTB/POZ or zinc finger domain, such as K4, K8 and K9 (Fig. 1), caused a loss of no more than 10% of the cells. Constructs expressing a complete or nearly complete POZ/BTB, such as K1, K2, K3 and K7 (Fig. 1), caused death of 60 to 95% of the cells. K3 and K7 fragments contain both SA domains, and the surviving fraction of cells overexpressing these fragments showed during mitosis multiple, strong and aberrant dots at putative centrosomal regions (Fig. 9). Aberrant distribution of the chromosomes (Fig. 9, DAPI stain) was accompanied with these abnormally shaped structures.

Bottom Line: In the present study we monitored Kaiso's subcellular localization during the cell cycle and found the following: (1) during interphase, Kaiso is located not only in the nucleus, but also on microtubular structures, including the centrosome; (2) at metaphase, it is present at the centrosomes and on the spindle microtubules; (3) during telophase, it accumulates at the midbody.We found that Kaiso is a genuine PCM component that belongs to a pericentrin molecular complex.Knockdown of Kaiso accelerated cell proliferation.

View Article: PubMed Central - PubMed

Affiliation: Department for Molecular Biomedical Research, VIB, Ghent, Belgium.

ABSTRACT
Kaiso is a BTB/POZ zinc finger protein known as a transcriptional repressor. It was originally identified through its in vitro association with the Armadillo protein p120ctn. Subcellular localization of Kaiso in cell lines and in normal and cancerous human tissues revealed that its expression is not restricted to the nucleus. In the present study we monitored Kaiso's subcellular localization during the cell cycle and found the following: (1) during interphase, Kaiso is located not only in the nucleus, but also on microtubular structures, including the centrosome; (2) at metaphase, it is present at the centrosomes and on the spindle microtubules; (3) during telophase, it accumulates at the midbody. We found that Kaiso is a genuine PCM component that belongs to a pericentrin molecular complex. We analyzed the functions of different domains of Kaiso by visualizing the subcellular distribution of GFP-tagged Kaiso fragments throughout the cell cycle. Our results indicate that two domains are responsible for targeting Kaiso to the centrosomes and microtubules. The first domain, designated SA1 for spindle-associated domain 1, is located in the center of the Kaiso protein and localizes at the spindle microtubules and centrosomes; the second domain, SA2, is an evolutionarily conserved domain situated just before the zinc finger domain and might be responsible for localizing Kaiso towards the centrosomal region. Constructs containing both SA domains and Kaiso's aminoterminal BTB/POZ domain triggered the formation of abnormal centrosomes. We also observed that overexpression of longer or full-length Kaiso constructs led to mitotic cell arrest and frequent cell death. Knockdown of Kaiso accelerated cell proliferation. Our data reveal a new target for Kaiso at the centrosomes and spindle microtubules during mitosis. They also strongly imply that Kaiso's function as a transcriptional regulator might be linked to the control of the cell cycle and to cell proliferation in cancer.

Show MeSH
Related in: MedlinePlus