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Aurora-A kinase is required for centrosome maturation in Caenorhabditis elegans.

Hannak E, Kirkham M, Hyman AA, Oegema K - J. Cell Biol. (2001)

Bottom Line: Consistent with this hypothesis, we find that AIR-1 is required for the increase in centrosomal gamma-tubulin and two other PCM components, ZYG-9 and CeGrip, as embryos enter mitosis.Furthermore, the AIR-1-dependent increase in centrosomal gamma-tubulin does not require MTs.These results suggest that aurora-A kinases are required to execute a MT-independent pathway for the recruitment of PCM during centrosome maturation.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute of Molecular Cell Biology and Genetics, 01307 Dresden, Germany.

ABSTRACT
Centrosomes mature as cells enter mitosis, accumulating gamma-tubulin and other pericentriolar material (PCM) components. This occurs concomitant with an increase in the number of centrosomally organized microtubules (MTs). Here, we use RNA-mediated interference (RNAi) to examine the role of the aurora-A kinase, AIR-1, during centrosome maturation in Caenorhabditis elegans. In air-1(RNAi) embryos, centrosomes separate normally, an event that occurs before maturation in C. elegans. After nuclear envelope breakdown, the separated centrosomes collapse together, and spindle assembly fails. In mitotic air-1(RNAi) embryos, centrosomal alpha-tubulin fluorescence intensity accumulates to only 40% of wild-type levels, suggesting a defect in the maturation process. Consistent with this hypothesis, we find that AIR-1 is required for the increase in centrosomal gamma-tubulin and two other PCM components, ZYG-9 and CeGrip, as embryos enter mitosis. Furthermore, the AIR-1-dependent increase in centrosomal gamma-tubulin does not require MTs. These results suggest that aurora-A kinases are required to execute a MT-independent pathway for the recruitment of PCM during centrosome maturation.

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AIR-1 is required for the accumulation of γ-tubulin, CeGrip, and ZYG-9 during centrosome maturation. Wild-type and air-1(RNAi) embryos stained for MTs and DNA (left, green and red) and for either γ-tubulin (A), CeGrip (B), or ZYG-9 (C) are shown. Inset in A is magnified 5.5-fold. Bar, 10 μm.
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fig4: AIR-1 is required for the accumulation of γ-tubulin, CeGrip, and ZYG-9 during centrosome maturation. Wild-type and air-1(RNAi) embryos stained for MTs and DNA (left, green and red) and for either γ-tubulin (A), CeGrip (B), or ZYG-9 (C) are shown. Inset in A is magnified 5.5-fold. Bar, 10 μm.

Mentions: The results above showed that a GFP–γ-tubulin fusion protein fails to accumulate at mitotic centrosomes in air-1(RNAi) embryos. To confirm this for endogenous γ-tubulin, we fixed air-1(RNAi) embryos and stained them for DNA, MTs, γ-tubulin, and AIR-1 (Fig. 4 A). In mitotic air-1(RNAi) embryos, AIR-1 was depleted to undetectable levels (Fig. 1 D), and γ-tubulin staining foci were much smaller than in wild-type (Fig. 4 A), consistent with the live analysis. Interestingly, each centrosome in the air-1(RNAi) embryos often appeared as a pair of small dots of γ-tubulin staining (Fig. 4 A, bottom right, inset). These dots are likely to be γ-tubulin associated with paired centrioles that have not yet separated.


Aurora-A kinase is required for centrosome maturation in Caenorhabditis elegans.

Hannak E, Kirkham M, Hyman AA, Oegema K - J. Cell Biol. (2001)

AIR-1 is required for the accumulation of γ-tubulin, CeGrip, and ZYG-9 during centrosome maturation. Wild-type and air-1(RNAi) embryos stained for MTs and DNA (left, green and red) and for either γ-tubulin (A), CeGrip (B), or ZYG-9 (C) are shown. Inset in A is magnified 5.5-fold. Bar, 10 μm.
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Related In: Results  -  Collection

Show All Figures
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fig4: AIR-1 is required for the accumulation of γ-tubulin, CeGrip, and ZYG-9 during centrosome maturation. Wild-type and air-1(RNAi) embryos stained for MTs and DNA (left, green and red) and for either γ-tubulin (A), CeGrip (B), or ZYG-9 (C) are shown. Inset in A is magnified 5.5-fold. Bar, 10 μm.
Mentions: The results above showed that a GFP–γ-tubulin fusion protein fails to accumulate at mitotic centrosomes in air-1(RNAi) embryos. To confirm this for endogenous γ-tubulin, we fixed air-1(RNAi) embryos and stained them for DNA, MTs, γ-tubulin, and AIR-1 (Fig. 4 A). In mitotic air-1(RNAi) embryos, AIR-1 was depleted to undetectable levels (Fig. 1 D), and γ-tubulin staining foci were much smaller than in wild-type (Fig. 4 A), consistent with the live analysis. Interestingly, each centrosome in the air-1(RNAi) embryos often appeared as a pair of small dots of γ-tubulin staining (Fig. 4 A, bottom right, inset). These dots are likely to be γ-tubulin associated with paired centrioles that have not yet separated.

Bottom Line: Consistent with this hypothesis, we find that AIR-1 is required for the increase in centrosomal gamma-tubulin and two other PCM components, ZYG-9 and CeGrip, as embryos enter mitosis.Furthermore, the AIR-1-dependent increase in centrosomal gamma-tubulin does not require MTs.These results suggest that aurora-A kinases are required to execute a MT-independent pathway for the recruitment of PCM during centrosome maturation.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute of Molecular Cell Biology and Genetics, 01307 Dresden, Germany.

ABSTRACT
Centrosomes mature as cells enter mitosis, accumulating gamma-tubulin and other pericentriolar material (PCM) components. This occurs concomitant with an increase in the number of centrosomally organized microtubules (MTs). Here, we use RNA-mediated interference (RNAi) to examine the role of the aurora-A kinase, AIR-1, during centrosome maturation in Caenorhabditis elegans. In air-1(RNAi) embryos, centrosomes separate normally, an event that occurs before maturation in C. elegans. After nuclear envelope breakdown, the separated centrosomes collapse together, and spindle assembly fails. In mitotic air-1(RNAi) embryos, centrosomal alpha-tubulin fluorescence intensity accumulates to only 40% of wild-type levels, suggesting a defect in the maturation process. Consistent with this hypothesis, we find that AIR-1 is required for the increase in centrosomal gamma-tubulin and two other PCM components, ZYG-9 and CeGrip, as embryos enter mitosis. Furthermore, the AIR-1-dependent increase in centrosomal gamma-tubulin does not require MTs. These results suggest that aurora-A kinases are required to execute a MT-independent pathway for the recruitment of PCM during centrosome maturation.

Show MeSH