Limits...
Stu2 promotes mitotic spindle elongation in anaphase.

Severin F, Habermann B, Huffaker T, Hyman T - J. Cell Biol. (2001)

Bottom Line: We further show that the activity of Stu2 is opposed by the activity of the kinesin-related protein Kip3.Reexamination of the kinesin homology tree suggests that KIP3 is the S. cerevisiae orthologue of the microtubule-destabilizing subfamily of kinesins (Kin I).We conclude that a balance of activity between evolutionally conserved microtubule-stabilizing and microtubule-destabilizing factors is essential for correct spindle elongation during anaphase B.

View Article: PubMed Central - PubMed

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

ABSTRACT
During anaphase, mitotic spindles elongate up to five times their metaphase length. This process, known as anaphase B, is essential for correct segregation of chromosomes. Here, we examine the control of spindle length during anaphase in the budding yeast Saccharomyces cerevisiae. We show that microtubule stabilization during anaphase requires the microtubule-associated protein Stu2. We further show that the activity of Stu2 is opposed by the activity of the kinesin-related protein Kip3. Reexamination of the kinesin homology tree suggests that KIP3 is the S. cerevisiae orthologue of the microtubule-destabilizing subfamily of kinesins (Kin I). We conclude that a balance of activity between evolutionally conserved microtubule-stabilizing and microtubule-destabilizing factors is essential for correct spindle elongation during anaphase B.

Show MeSH
Unrooted phylogenetic tree of the kinesin motor domains. A multiple sequence alignment for the kinesin motor domain sequences was constructed using the program ClustalX, a bootstrapped phylogenetic tree (neighbor-joining tree), was calculated using the same program and displayed as a radial tree using the program TreeView. Congruent with biological data, the Kip3 subfamily is found on the same branch as the MCAK/Kif2 subfamily, suggesting close evolutionary relationship between the members of these subfamilies. Only the bootstrap values for the Kip3 and MCAK/Kif2 subbranches are shown (538 and 528 for the branch leading to Kip2, Kip3, MACK/Kif2, and Kip3, MACK/Kif2, respectively).
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2169465&req=5

Figure 7: Unrooted phylogenetic tree of the kinesin motor domains. A multiple sequence alignment for the kinesin motor domain sequences was constructed using the program ClustalX, a bootstrapped phylogenetic tree (neighbor-joining tree), was calculated using the same program and displayed as a radial tree using the program TreeView. Congruent with biological data, the Kip3 subfamily is found on the same branch as the MCAK/Kif2 subfamily, suggesting close evolutionary relationship between the members of these subfamilies. Only the bootstrap values for the Kip3 and MCAK/Kif2 subbranches are shown (538 and 528 for the branch leading to Kip2, Kip3, MACK/Kif2, and Kip3, MACK/Kif2, respectively).

Mentions: In summary, we conclude that stabilization of the central spindle during anaphase B depends on a balance between the opposing activities of Stu2 and Kip3. Our data further suggests that KIP3 is the functional orthologue of the Kin I subfamily of proteins. Members of this family have been found in all organisms examined (Desai et al. 1999) except yeast. Our discovery of the effect of Kip3 in opposing Stu2 suggests either that the Kin I family evolved an orthologous function in metazoans, or that the bioinformatics analysis has missed the similarities in sequence. On the basis of our observations, we reanalyzed the relationship of XKCM1 to the kinesin-related proteins in yeast. We recalculated a phylogenetic tree from a multiple sequence alignment of the kinesin superfamily using only the sequence of the kinesin motor domain. In the resulting phylogenetic tree (Fig. 7), the Kip3 family of kinesin motor domain proteins did congregate with the MCAK/Kif2 (Kin I) subfamily, which is in accordance with our experimental data. The resulting tree is opposed to a recently published phylogenetic tree (available at kinesin homepage, http://www.proweb.org/kinesin/ KinesinTree.html), in which the two subfamilies are found on two separate branches of the phylogenetic tree. The observed disagreement is most likely due to the fact that the kinesin domain from Kip3 we used for building the multiple sequence alignment differs in length from the one at kinesin homepage. In the case of S. cerevisiae KIP3, the sequence was 97 amino acids longer than the one used for the tree at the kinesin homepage. This results in a 56–amino acid gap produced by ScKip3, yet the conservation at the very NH2 terminus of the kinesin domains, which includes a V-x-[V,I]-R-x-R-P-[V,I,L,F]-x(3)-[E,N,Q,D] motif in this part of the kinesin domain that is present in most of the family members, justifies the inclusion of this part of Kip3 family members. This motif in the first conserved part of the kinesin domain furthermore aligns to a conserved β-sheet (β-1) (data not shown). In addition, we excluded gap columns, and only conserved alignment blocks were used for calculating the phylogenetic tree. Sequences that produced gaps in otherwise conserved alignment blocks were excluded from the analysis. Two additional phylogenetic trees have been published before (Goodson et al. 1994; Moore and Endow 1996), none of which contain the Kip3 subfamily of kinesin motor domain proteins and therefore make comparison to our results impossible. Interestingly, MCAK, a member of Kin I subfamily of kinesins, is localized to kinetochores in mammalian cells (Wordeman and Mitchison 1995). Since it now appears to be part of this subfamily, perhaps Kip3 is a kinetochore component in S. cerevisiae as well, thus explaining the chromosome missegregation.


Stu2 promotes mitotic spindle elongation in anaphase.

Severin F, Habermann B, Huffaker T, Hyman T - J. Cell Biol. (2001)

Unrooted phylogenetic tree of the kinesin motor domains. A multiple sequence alignment for the kinesin motor domain sequences was constructed using the program ClustalX, a bootstrapped phylogenetic tree (neighbor-joining tree), was calculated using the same program and displayed as a radial tree using the program TreeView. Congruent with biological data, the Kip3 subfamily is found on the same branch as the MCAK/Kif2 subfamily, suggesting close evolutionary relationship between the members of these subfamilies. Only the bootstrap values for the Kip3 and MCAK/Kif2 subbranches are shown (538 and 528 for the branch leading to Kip2, Kip3, MACK/Kif2, and Kip3, MACK/Kif2, respectively).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2169465&req=5

Figure 7: Unrooted phylogenetic tree of the kinesin motor domains. A multiple sequence alignment for the kinesin motor domain sequences was constructed using the program ClustalX, a bootstrapped phylogenetic tree (neighbor-joining tree), was calculated using the same program and displayed as a radial tree using the program TreeView. Congruent with biological data, the Kip3 subfamily is found on the same branch as the MCAK/Kif2 subfamily, suggesting close evolutionary relationship between the members of these subfamilies. Only the bootstrap values for the Kip3 and MCAK/Kif2 subbranches are shown (538 and 528 for the branch leading to Kip2, Kip3, MACK/Kif2, and Kip3, MACK/Kif2, respectively).
Mentions: In summary, we conclude that stabilization of the central spindle during anaphase B depends on a balance between the opposing activities of Stu2 and Kip3. Our data further suggests that KIP3 is the functional orthologue of the Kin I subfamily of proteins. Members of this family have been found in all organisms examined (Desai et al. 1999) except yeast. Our discovery of the effect of Kip3 in opposing Stu2 suggests either that the Kin I family evolved an orthologous function in metazoans, or that the bioinformatics analysis has missed the similarities in sequence. On the basis of our observations, we reanalyzed the relationship of XKCM1 to the kinesin-related proteins in yeast. We recalculated a phylogenetic tree from a multiple sequence alignment of the kinesin superfamily using only the sequence of the kinesin motor domain. In the resulting phylogenetic tree (Fig. 7), the Kip3 family of kinesin motor domain proteins did congregate with the MCAK/Kif2 (Kin I) subfamily, which is in accordance with our experimental data. The resulting tree is opposed to a recently published phylogenetic tree (available at kinesin homepage, http://www.proweb.org/kinesin/ KinesinTree.html), in which the two subfamilies are found on two separate branches of the phylogenetic tree. The observed disagreement is most likely due to the fact that the kinesin domain from Kip3 we used for building the multiple sequence alignment differs in length from the one at kinesin homepage. In the case of S. cerevisiae KIP3, the sequence was 97 amino acids longer than the one used for the tree at the kinesin homepage. This results in a 56–amino acid gap produced by ScKip3, yet the conservation at the very NH2 terminus of the kinesin domains, which includes a V-x-[V,I]-R-x-R-P-[V,I,L,F]-x(3)-[E,N,Q,D] motif in this part of the kinesin domain that is present in most of the family members, justifies the inclusion of this part of Kip3 family members. This motif in the first conserved part of the kinesin domain furthermore aligns to a conserved β-sheet (β-1) (data not shown). In addition, we excluded gap columns, and only conserved alignment blocks were used for calculating the phylogenetic tree. Sequences that produced gaps in otherwise conserved alignment blocks were excluded from the analysis. Two additional phylogenetic trees have been published before (Goodson et al. 1994; Moore and Endow 1996), none of which contain the Kip3 subfamily of kinesin motor domain proteins and therefore make comparison to our results impossible. Interestingly, MCAK, a member of Kin I subfamily of kinesins, is localized to kinetochores in mammalian cells (Wordeman and Mitchison 1995). Since it now appears to be part of this subfamily, perhaps Kip3 is a kinetochore component in S. cerevisiae as well, thus explaining the chromosome missegregation.

Bottom Line: We further show that the activity of Stu2 is opposed by the activity of the kinesin-related protein Kip3.Reexamination of the kinesin homology tree suggests that KIP3 is the S. cerevisiae orthologue of the microtubule-destabilizing subfamily of kinesins (Kin I).We conclude that a balance of activity between evolutionally conserved microtubule-stabilizing and microtubule-destabilizing factors is essential for correct spindle elongation during anaphase B.

View Article: PubMed Central - PubMed

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

ABSTRACT
During anaphase, mitotic spindles elongate up to five times their metaphase length. This process, known as anaphase B, is essential for correct segregation of chromosomes. Here, we examine the control of spindle length during anaphase in the budding yeast Saccharomyces cerevisiae. We show that microtubule stabilization during anaphase requires the microtubule-associated protein Stu2. We further show that the activity of Stu2 is opposed by the activity of the kinesin-related protein Kip3. Reexamination of the kinesin homology tree suggests that KIP3 is the S. cerevisiae orthologue of the microtubule-destabilizing subfamily of kinesins (Kin I). We conclude that a balance of activity between evolutionally conserved microtubule-stabilizing and microtubule-destabilizing factors is essential for correct spindle elongation during anaphase B.

Show MeSH