Molecular dissection of condensin II-mediated chromosome assembly using in vitro assays
- RIKEN, Japan
Abstract
In vertebrates, condensin I and condensin II cooperate to assemble rod-shaped chromosomes during mitosis. Although the mechanism of action and regulation of condensin I have been studied extensively, our corresponding knowledge of condensin II remains very limited. By introducing recombinant condensin II complexes into Xenopus egg extracts, we dissect the roles of its individual subunits in chromosome assembly. We find that one of two HEAT subunits, CAP-D3, plays a crucial role in condensin II-mediated assembly of chromosome axes whereas the other HEAT subunit, CAP-G2, has a very strong negative impact on this process. The SMC ATPase and the basic amino acid clusters of the kleisin subunit CAP-H2 are essential for this process. Deletion of the C-terminal tail of CAP-D3 increases the ability of condensin II to assemble chromosomes and further exposes a hidden function of CAP-G2 in the lateral compaction of chromosomes. Taken together, our results uncover a multilayered regulatory mechanism unique to condensin II, and provide profound implications for the evolution of condensin II.
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All data generated or analyzed during this experimental study are included in the manuscript as source data.
Article and author information
Author details
Kazuhisa Kinoshita
Funding
Japan Society for the Promotion of Science (#20K15723)
- Makoto M Yoshida
Japan Society for the Promotion of Science (#15K06959)
- Kazuhisa Kinoshita
Japan Society for the Promotion of Science (#19K06499)
- Kazuhisa Kinoshita
Japan Society for the Promotion of Science (#18H02381)
- Keishi Shintomi
Japan Society for the Promotion of Science (#19H05755)
- Keishi Shintomi
Japan Society for the Promotion of Science (#18H05276)
- Tatsuya Hirano
Japan Society for the Promotion of Science (#20H0593)
- Tatsuya Hirano
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Animal experimentation: Female Xenopus laevis frogs (RRID: NXR 0.031, Hamamatsu Seibutsu-Kyozai) were used to lay eggs to harvest Xenopus egg extract (Hirano et al., 1997). Male X. laevis frogs (RRID: NXR 0.031, Hamamatsu Seibutsu-Kyozai) were dissected to prepare sperm nuclei from testes (Shintomi and Hirano, 2017). Frogs were used in compliance with the institutional regulations of the RIKEN Wako Campus. Mice (BALB/c × C57BL/6J)F1) for sperm nuclei (Shintomi et al., 2017) were used in compliance with protocols approved by the Animal Care and Use Committee of the University of Tokyo (for M. Ohsugi who provided mouse sperm).
Copyright
© 2022, Yoshida et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Molecular dissection of condensin II-mediated chromosome assembly using in vitro assays
eLife 11:e78984.
https://doi.org/10.7554/eLife.78984
