Mechanisms ensuring proper chromosome segregation in mitosis
To maintain genetic integrity, eukaryotic cells must segregate their duplicated chromosomes to their daughter cells with high fidelity during mitosis. The unravelling of the mechanisms ensuring proper chromosome segregation should improve our understanding of various human diseases such as cancers and congenital disorders, which are characterized by chromosome instability and aneuploidy.
We study mechanisms of chromosome segregation in budding yeast and human cells, taking advantages of these systems. Basic mechanisms of chromosome regulation are well conserved from yeast to humans. With yeast cells, we can flexibly set up experiments and quickly get results, whereas with human cells, we can obtain information more directly relevant to human health and diseases. We are currently focusing on the following research projects:
1) Kinetochore-microtubule interaction in mitosis
Kinetochores are large protein complexes formed at the centromere regions of chromosomes. For high-fidelity chromosome segregation, kinetochores must be properly caught on the mitotic spindle (reviewed in Tanaka TU 2010). We have found that kinetochores initially interact with the lateral surface of a single microtubule extending from a spindle pole (Figure 1, step 2; Tanaka K et al, 2005, Kitamura et al, 2007); this process is often facilitated by microtubules generated at kinetochores (Figure 1, step 1; Kitamura et al, 2010; Vasileva et al 2017). Subsequently kinetochores are tethered at the microtubule plus ends (Figure 1, step 3; Figure 2; Tanaka K et al, 2007; Maure et al, 2011; Kalantzaki et al 2015). When this process fails, failsafe mechanisms prevent kinetochore detachment from a microtubule (Figure 2; Gandhi et al, 2011; Yue et al 2017). Moreover we found that, in human cells, contractile acto-myosin network on nuclear envelope remnants reduces chromosome scattering and facilitates kinetochore–microtubule interaction in early mitosis (Figure 3; Booth et al 2019).
Following the initial interaction with microtubules, sister kinetochores must interact with microtubules extending from opposite spindle poles (chromosome bi-orientation) before anaphase onset (reviewed in Doodhi and Tanaka TU 2022; Tanaka TU and Zhang 2022). If this interaction occurs with aberrant orientation (Figure 1, step 4), such errors must be corrected by changeover of the kinetochore–microtubule attachment (error correction; Figure 2, step 5). We have found that Aurora B/Ipl1 kinase and other factors essentially promote this process, using live-cell imaging (Tanaka TU et al 2002; Dewar et al, 2004; Maure et al, 2007; Kalantzaki et al 2015) and in vitro reconstitution of kinetochore–microtubule interface (Doodhi et al, 2021) (Figure 4); in particular, Aurora B/Ipl1 at centromeres/inner kinetochores plays a central role (Garcia-Rodriguez et al 2019). Once bi-orientation is established and tension is applied across sister kinetochores, kinetochore–microtubule interaction is stabilized (Figure 1, step 6; Keating et al, 2009). Sister chromatid cohesion around centromeres plays important roles in this process (Natsume et al, 2013; reviewed in Tanaka TU et al, 2013). We are investigating these mechanisms further in yeast and human cells.
We study mechanisms of chromosome segregation in budding yeast and human cells, taking advantages of these systems. Basic mechanisms of chromosome regulation are well conserved from yeast to humans. With yeast cells, we can flexibly set up experiments and quickly get results, whereas with human cells, we can obtain information more directly relevant to human health and diseases. We are currently focusing on the following research projects:
1) Kinetochore-microtubule interaction in mitosis
Kinetochores are large protein complexes formed at the centromere regions of chromosomes. For high-fidelity chromosome segregation, kinetochores must be properly caught on the mitotic spindle (reviewed in Tanaka TU 2010). We have found that kinetochores initially interact with the lateral surface of a single microtubule extending from a spindle pole (Figure 1, step 2; Tanaka K et al, 2005, Kitamura et al, 2007); this process is often facilitated by microtubules generated at kinetochores (Figure 1, step 1; Kitamura et al, 2010; Vasileva et al 2017). Subsequently kinetochores are tethered at the microtubule plus ends (Figure 1, step 3; Figure 2; Tanaka K et al, 2007; Maure et al, 2011; Kalantzaki et al 2015). When this process fails, failsafe mechanisms prevent kinetochore detachment from a microtubule (Figure 2; Gandhi et al, 2011; Yue et al 2017). Moreover we found that, in human cells, contractile acto-myosin network on nuclear envelope remnants reduces chromosome scattering and facilitates kinetochore–microtubule interaction in early mitosis (Figure 3; Booth et al 2019).
Following the initial interaction with microtubules, sister kinetochores must interact with microtubules extending from opposite spindle poles (chromosome bi-orientation) before anaphase onset (reviewed in Doodhi and Tanaka TU 2022; Tanaka TU and Zhang 2022). If this interaction occurs with aberrant orientation (Figure 1, step 4), such errors must be corrected by changeover of the kinetochore–microtubule attachment (error correction; Figure 2, step 5). We have found that Aurora B/Ipl1 kinase and other factors essentially promote this process, using live-cell imaging (Tanaka TU et al 2002; Dewar et al, 2004; Maure et al, 2007; Kalantzaki et al 2015) and in vitro reconstitution of kinetochore–microtubule interface (Doodhi et al, 2021) (Figure 4); in particular, Aurora B/Ipl1 at centromeres/inner kinetochores plays a central role (Garcia-Rodriguez et al 2019). Once bi-orientation is established and tension is applied across sister kinetochores, kinetochore–microtubule interaction is stabilized (Figure 1, step 6; Keating et al, 2009). Sister chromatid cohesion around centromeres plays important roles in this process (Natsume et al, 2013; reviewed in Tanaka TU et al, 2013). We are investigating these mechanisms further in yeast and human cells.
Figure 3. During prophase in human cells, the LINC complex promotes the accumulation of acto-myosin on the nuclear envelope (i, ii). Upon the nuclear envelope breakdown, myosin II promotes contraction of the acto-myosin network (iii), which reduces chromosome scattering and facilitates correct chromosome interaction with the mitotic spindle (iv) (Booth et al 2019).
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2) Sister chromatid cohesion and chromosome compaction
Duplicated chromosomes (sister chromatids) are held together by sister chromatid cohesion until chromosome segregation takes place in anaphase. Without this cohesion, cells cannot distinguish a pair of sister chromatids destined for segregation. In particular, robust sister chromatid cohesion around kinetochores is crucial for this purpose. In budding yeast, we found that this robust cohesion is facilitated by Dbf4-dependent kinase recruited to kinetochores (Natsume et al, 2013). Sister chromatid cohesion is maintained until metaphase, but it must be quickly removed from chromosomes upon anaphase onset. We have found novel mechanisms that facilitate this removal (Figure 5: Li et al, 2017; Renshaw et al 2010) independently of a well-known separase-dependent mechanism. We are currently studying these mechanisms further.
When human cells enter mitosis, sister chromatids must be resolved and compacted to prepare for chromosome segregation. We have developed a novel fluorescence reporter system to analyse how these processes proceed and are coordinated (Eykelenboom et al 2019; reviewed in Eykelenboom and Tanaka 2020). We found novel roles of cohesin regulators and condensin I and II in this, and study relevant mechanisms further using this system.
Duplicated chromosomes (sister chromatids) are held together by sister chromatid cohesion until chromosome segregation takes place in anaphase. Without this cohesion, cells cannot distinguish a pair of sister chromatids destined for segregation. In particular, robust sister chromatid cohesion around kinetochores is crucial for this purpose. In budding yeast, we found that this robust cohesion is facilitated by Dbf4-dependent kinase recruited to kinetochores (Natsume et al, 2013). Sister chromatid cohesion is maintained until metaphase, but it must be quickly removed from chromosomes upon anaphase onset. We have found novel mechanisms that facilitate this removal (Figure 5: Li et al, 2017; Renshaw et al 2010) independently of a well-known separase-dependent mechanism. We are currently studying these mechanisms further.
When human cells enter mitosis, sister chromatids must be resolved and compacted to prepare for chromosome segregation. We have developed a novel fluorescence reporter system to analyse how these processes proceed and are coordinated (Eykelenboom et al 2019; reviewed in Eykelenboom and Tanaka 2020). We found novel roles of cohesin regulators and condensin I and II in this, and study relevant mechanisms further using this system.
Figure 6. tet and lac operator arrays were inserted 245 kp apart in the middle of the right arm of human chromosome 5. The arrays were visualized as red and green fluorescent dots in cells. By observing how their patterns change, we were able to analyse how sister chromatid resolution and compaction proceed from late G2 phase to early mitosis (Eykelenboom et al 2019).
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3) Other related topics
Before chromosome segregation takes place, chromosomes must be duplicated properly. We have been studying how chromosome duplication is regulated in space and time within the nucleus in budding yeast. We found that how replication factories are formed to organize DNA replication (Kitamura et al, 2006; Saner et al, 2013) and how replication timing is regulated around centromeres (Natsume et al, 2013). Meanwhile, we have also studied evolution of the centromere. We discovered a novel type of centromere in budding yeast, which sheds new light on the centromere evolution in the yeast lineage (Kobayashi et al, 2015).
Before chromosome segregation takes place, chromosomes must be duplicated properly. We have been studying how chromosome duplication is regulated in space and time within the nucleus in budding yeast. We found that how replication factories are formed to organize DNA replication (Kitamura et al, 2006; Saner et al, 2013) and how replication timing is regulated around centromeres (Natsume et al, 2013). Meanwhile, we have also studied evolution of the centromere. We discovered a novel type of centromere in budding yeast, which sheds new light on the centromere evolution in the yeast lineage (Kobayashi et al, 2015).
Publications
Tanaka TU & Zhang T. SWAP, SWITCH and STABILIZE: mechanisms of kinetochore–microtubule error correction. Cells, 11, 1462 (2022). [Pubmed] [PDF]
Doodhi H & Tanaka TU. Swap and stop – kinetochores play error correction with microtubules, BioEssay, 2100246 (2022). [Pubmed] [PDF]
Doodhi H, Kasciukovic T, Clayton L & Tanaka TU. Aurora B switches relative strength of kinetochore–microtubule attachment modes for error correction. J. Cell Biol. 220, e202011117 (2021). [Pubmed] [PDF]
Eykelenboom J & Tanaka TU. Zoom in on chromosome dynamics. Cell Cycle 19, 1422-32 (2020). [Pubmed] [PDF]
Booth AJR., Yue, Z, Eykelenboom JK, Luxton GWG, Hochegger H & Tanaka TU. Contractile actomyosin network on nuclear envelope remants positions chromososomes for mitosis. eLife 8, e46902 (2019). [Pubmed] [PDF]
Garcia-Rodriguez LJ, Kasciukovic T*, Denninger V* & Tanaka TU. Aurora B-INCENP localization at centromeres/inner kinetochores is essential for chromosome bi-orientation in budding yeast. Curr Biol, 29, 1536-44 (2019). (* equal contribution) [Pubmed] [PDF]
Eykelenboom, J.K., Gierlinski, M.*, Yue, Z.*, Hegarat, N.*, Pollard, H., Fukagawa, T., Hochegger, H., & Tanaka, T.U. Live imaging of marked chromosome regions reveals their dynamic resolution and compaction in mitosis. J. Cell Biol. 218, 1531-52 (2019). (* equal contribution) [Pubmed] [PDF]
Li S, Yue Z & Tanaka TU. Smc3 deacetylation by Hos1 facilitates efficient dissolution of sister chromatid cohesion during early anaphase. Mol Cell. 68, 605-14 (2017). [Pubmed] [PDF]
Yue Z, Komoto S, Gierlinski M, Pasquali D, Kitamura E & Tanaka TU. Mechanisms mitigating problems associated with multiple kinetochores on one microtubule in early mitosis. J. Cell Sci, 130, 2266-76. (2017). [Pubmed] [PDF]
Vasileva V, Gierlinski M, Yue Z, O’Reilly N, Kitamura E & Tanaka TU. Molecular mechanisms facilitating the initial kinetochore encounter with spindle microtubules. J Cell Biol, 216, 1609-22. (2017). [Pubmed] [PDF]
Kalantzaki M, Kitamura E, Zhang T, Mino A, Novák B & Tanaka TU. Kinetochore–microtubule error correction is driven by differentially regulated interaction modes. Nat Cell Biol, 17, 421-33, (2015). [Pubmed] [PDF] [PMC-PDF]
Kobayashi N, Suzuki Y, Schoenfeld LW, Müller CA, Nieduszynski C, Wolfe KH & Tanaka TU. Discovery of an unconventional centromere in budding yeast redefines evolution of point centromeres. Curr Biol, 25, 2026-33. (2015). [Pubmed] [PDF]
Tanaka TU, Clayton L & Natsume T. Three wise centromere functions: see no error, hear no break, speak no delay. EMBO Rep, 14, 1073-83. (2013). [Pubmed] [PDF]
Natsume T, Müller CA, Katou Y, Retkute R, Gierlinski M, Araki H, Blow JJ, Shirahige, K, Nieduszynski CA & Tanaka TU. Kinetochores coordinate pericentromeric cohesion and early DNA replication by Cdc7-Dbf4 kinase recruitment. Mol Cell, 50, 661-74. (2013). [Pubmed] [PDF]
Saner N, Karschau J, Natsume T, Gierlinski M, Retkute R, Hawkins M, Nieduszynski CA, Blow JJ, de Moura APS & Tanaka TU. Stochastic association of neighboring replicons creates replication factories in budding yeast. J Cell Biol, 202, 1001-12. (2013). [Pubmed] [PDF]
Gandhi SR, Gierlinski M, Mino A, Tanaka K, Kitamura E, Clayton L & Tanaka TU. Kinetochore-dependent microtubule rescue ensures their efficient and sustained interactions in early mitosis. Dev Cell, 21, 920-33. (2011). [Pubmed] [PDF]
Maure J-F*, Komoto S*, Oku Y, Mino A, Pasqualato S, Natsume K, Clayton L, Musacchio A & Tanaka TU. The Ndc80 loop region facilitates formation of kinetochore attachment to the dynamic microtubule plus end. Curr Biol, 21, 207-13. (2011). (* equal contribution) [Pubmed] [PDF]
Tanaka TU. Kinetochore-microtubule interactions: steps towards bi-orientation (EMBO members’ review). EMBO J, 29, 4070-82. (2010). [Pubmed] [PDF]
Renshaw MJ, Ward JJ, Kanemaki M, Natsume K, Nedelec FJ & Tanaka TU. Condensins promote chromosome recoiling during early anaphase to complete sister chromatid separation. Dev Cell, 19, 232-44. (2010). [Pubmed] [PDF]
Kitamura E*, Tanaka K*, Komoto S*, Kitamura Y, Antony C & Tanaka TU. Kinetochores generate microtubules with distal plus ends: their roles and limited lifetime in mitosis. Dev Cell, 18, 248-59. (2010). (* equal contribution) [Pubmed] [PDF]
Keating P, Rachidi N, Tanaka TU* & Stark MJR*. Ipl1-dependent phosphorylation of Dam1 is reduced by tension applied on kinetochores. J Cell Sci,122, 4375-82. (2009). (* corresponding authors) [Pubmed] [PDF]
Maure J-F, Kitamura E & Tanaka TU. Mps1 kinase promotes sister kinetochore bi-orientation by a tension-dependent mechanism. Curr Biol, 17, 2175-82. (2007). [Pubmed] [PDF]
Kitamura E, Tanaka K, Kitamura Y & Tanaka TU. Kinetochore-microtubule interaction during S phase in Saccharomyces cerevisiae. Genes Dev, 21, 3319-30. (2007). [Pubmed] [PDF]
Tanaka K, Kitamura E, Kitamura Y & Tanaka TU. Molecular mechanisms of microtubule-dependent kinetochore transport towards spindle poles. J Cell Biol, 178, 269-81. (2007). [Pubmed] [PDF]
Kitamura E, Blow JJ & Tanaka TU. Live-cell imaging reveals replication of individual replicons in eukaryotic replication factories. Cell, 125, 1297-308. (2006). [Pubmed] [PDF]
Tanaka K, Mukae N, Dewar H, van Breugel M, James EK, Prescott AR, Antony C & Tanaka TU. Molecular mechanisms of kinetochore capture by spindle microtubules. Nature 434, 987-94. (2005). [Pubmed] [PDF]
Dewar H, Tanaka K, Nasmyth K & Tanaka TU. Tension between two kinetochores suffices for their bi-orientation on the mitotic spindle. Nature 428, 93-97. (2004). [Pubmed] [PDF]
Tanaka TU*, Rachidi N, Janke C, Pereira G, Galova M, Schiebel E, Stark MJ & Nasmyth K. Evidence that the Ipl1-Sli15 (Aurora kinase-INCENP) complex promotes chromosome bi-orientation by altering kinetochore-spindle pole connections. Cell 108, 317-29. (2002). (*corresponding author) [Pubmed] [PDF]
Tanaka TU & Zhang T. SWAP, SWITCH and STABILIZE: mechanisms of kinetochore–microtubule error correction. Cells, 11, 1462 (2022). [Pubmed] [PDF]
Doodhi H & Tanaka TU. Swap and stop – kinetochores play error correction with microtubules, BioEssay, 2100246 (2022). [Pubmed] [PDF]
Doodhi H, Kasciukovic T, Clayton L & Tanaka TU. Aurora B switches relative strength of kinetochore–microtubule attachment modes for error correction. J. Cell Biol. 220, e202011117 (2021). [Pubmed] [PDF]
Eykelenboom J & Tanaka TU. Zoom in on chromosome dynamics. Cell Cycle 19, 1422-32 (2020). [Pubmed] [PDF]
Booth AJR., Yue, Z, Eykelenboom JK, Luxton GWG, Hochegger H & Tanaka TU. Contractile actomyosin network on nuclear envelope remants positions chromososomes for mitosis. eLife 8, e46902 (2019). [Pubmed] [PDF]
Garcia-Rodriguez LJ, Kasciukovic T*, Denninger V* & Tanaka TU. Aurora B-INCENP localization at centromeres/inner kinetochores is essential for chromosome bi-orientation in budding yeast. Curr Biol, 29, 1536-44 (2019). (* equal contribution) [Pubmed] [PDF]
Eykelenboom, J.K., Gierlinski, M.*, Yue, Z.*, Hegarat, N.*, Pollard, H., Fukagawa, T., Hochegger, H., & Tanaka, T.U. Live imaging of marked chromosome regions reveals their dynamic resolution and compaction in mitosis. J. Cell Biol. 218, 1531-52 (2019). (* equal contribution) [Pubmed] [PDF]
Li S, Yue Z & Tanaka TU. Smc3 deacetylation by Hos1 facilitates efficient dissolution of sister chromatid cohesion during early anaphase. Mol Cell. 68, 605-14 (2017). [Pubmed] [PDF]
Yue Z, Komoto S, Gierlinski M, Pasquali D, Kitamura E & Tanaka TU. Mechanisms mitigating problems associated with multiple kinetochores on one microtubule in early mitosis. J. Cell Sci, 130, 2266-76. (2017). [Pubmed] [PDF]
Vasileva V, Gierlinski M, Yue Z, O’Reilly N, Kitamura E & Tanaka TU. Molecular mechanisms facilitating the initial kinetochore encounter with spindle microtubules. J Cell Biol, 216, 1609-22. (2017). [Pubmed] [PDF]
Kalantzaki M, Kitamura E, Zhang T, Mino A, Novák B & Tanaka TU. Kinetochore–microtubule error correction is driven by differentially regulated interaction modes. Nat Cell Biol, 17, 421-33, (2015). [Pubmed] [PDF] [PMC-PDF]
Kobayashi N, Suzuki Y, Schoenfeld LW, Müller CA, Nieduszynski C, Wolfe KH & Tanaka TU. Discovery of an unconventional centromere in budding yeast redefines evolution of point centromeres. Curr Biol, 25, 2026-33. (2015). [Pubmed] [PDF]
Tanaka TU, Clayton L & Natsume T. Three wise centromere functions: see no error, hear no break, speak no delay. EMBO Rep, 14, 1073-83. (2013). [Pubmed] [PDF]
Natsume T, Müller CA, Katou Y, Retkute R, Gierlinski M, Araki H, Blow JJ, Shirahige, K, Nieduszynski CA & Tanaka TU. Kinetochores coordinate pericentromeric cohesion and early DNA replication by Cdc7-Dbf4 kinase recruitment. Mol Cell, 50, 661-74. (2013). [Pubmed] [PDF]
Saner N, Karschau J, Natsume T, Gierlinski M, Retkute R, Hawkins M, Nieduszynski CA, Blow JJ, de Moura APS & Tanaka TU. Stochastic association of neighboring replicons creates replication factories in budding yeast. J Cell Biol, 202, 1001-12. (2013). [Pubmed] [PDF]
Gandhi SR, Gierlinski M, Mino A, Tanaka K, Kitamura E, Clayton L & Tanaka TU. Kinetochore-dependent microtubule rescue ensures their efficient and sustained interactions in early mitosis. Dev Cell, 21, 920-33. (2011). [Pubmed] [PDF]
Maure J-F*, Komoto S*, Oku Y, Mino A, Pasqualato S, Natsume K, Clayton L, Musacchio A & Tanaka TU. The Ndc80 loop region facilitates formation of kinetochore attachment to the dynamic microtubule plus end. Curr Biol, 21, 207-13. (2011). (* equal contribution) [Pubmed] [PDF]
Tanaka TU. Kinetochore-microtubule interactions: steps towards bi-orientation (EMBO members’ review). EMBO J, 29, 4070-82. (2010). [Pubmed] [PDF]
Renshaw MJ, Ward JJ, Kanemaki M, Natsume K, Nedelec FJ & Tanaka TU. Condensins promote chromosome recoiling during early anaphase to complete sister chromatid separation. Dev Cell, 19, 232-44. (2010). [Pubmed] [PDF]
Kitamura E*, Tanaka K*, Komoto S*, Kitamura Y, Antony C & Tanaka TU. Kinetochores generate microtubules with distal plus ends: their roles and limited lifetime in mitosis. Dev Cell, 18, 248-59. (2010). (* equal contribution) [Pubmed] [PDF]
Keating P, Rachidi N, Tanaka TU* & Stark MJR*. Ipl1-dependent phosphorylation of Dam1 is reduced by tension applied on kinetochores. J Cell Sci,122, 4375-82. (2009). (* corresponding authors) [Pubmed] [PDF]
Maure J-F, Kitamura E & Tanaka TU. Mps1 kinase promotes sister kinetochore bi-orientation by a tension-dependent mechanism. Curr Biol, 17, 2175-82. (2007). [Pubmed] [PDF]
Kitamura E, Tanaka K, Kitamura Y & Tanaka TU. Kinetochore-microtubule interaction during S phase in Saccharomyces cerevisiae. Genes Dev, 21, 3319-30. (2007). [Pubmed] [PDF]
Tanaka K, Kitamura E, Kitamura Y & Tanaka TU. Molecular mechanisms of microtubule-dependent kinetochore transport towards spindle poles. J Cell Biol, 178, 269-81. (2007). [Pubmed] [PDF]
Kitamura E, Blow JJ & Tanaka TU. Live-cell imaging reveals replication of individual replicons in eukaryotic replication factories. Cell, 125, 1297-308. (2006). [Pubmed] [PDF]
Tanaka K, Mukae N, Dewar H, van Breugel M, James EK, Prescott AR, Antony C & Tanaka TU. Molecular mechanisms of kinetochore capture by spindle microtubules. Nature 434, 987-94. (2005). [Pubmed] [PDF]
Dewar H, Tanaka K, Nasmyth K & Tanaka TU. Tension between two kinetochores suffices for their bi-orientation on the mitotic spindle. Nature 428, 93-97. (2004). [Pubmed] [PDF]
Tanaka TU*, Rachidi N, Janke C, Pereira G, Galova M, Schiebel E, Stark MJ & Nasmyth K. Evidence that the Ipl1-Sli15 (Aurora kinase-INCENP) complex promotes chromosome bi-orientation by altering kinetochore-spindle pole connections. Cell 108, 317-29. (2002). (*corresponding author) [Pubmed] [PDF]