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.

Data availability

All data generated or analyzed during this experimental study are included in the manuscript as source data.

Article and author information

Author details

  1. Makoto M Yoshida

    Chromosome Dynamics Laboratory, RIKEN, Wako, Japan
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0618-1717
  2. Kazuhisa Kinoshita

    Chromosome Dynamics Laboratory, RIKEN, Wako, Japan
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0882-4296
  3. Yuuki Aizawa

    Chromosome Dynamics Laboratory, RIKEN, Wako, Japan
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5002-7557
  4. Shoji Tane

    Chromosome Dynamics Laboratory, RIKEN, Wako, Japan
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0209-347X
  5. Daisuke Yamashita

    Chromosome Dynamics Laboratory, RIKEN, Wako, Japan
    Competing interests
    Daisuke Yamashita, is currently affiliated with Otsuka Pharmaceutical Co., Ltd. The author has no financial interests to declare..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6054-5617
  6. Keishi Shintomi

    Chromosome Dynamics Laboratory, RIKEN, Wako, Japan
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-0484-9901
  7. Tatsuya Hirano

    Chromosome Dynamics Laboratory, RIKEN, Wako, Japan
    For correspondence
    hiranot@riken.jp
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4219-6473

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.

Metrics

  • 1,785
    views
  • 433
    downloads
  • 13
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Makoto M Yoshida
  2. Kazuhisa Kinoshita
  3. Yuuki Aizawa
  4. Shoji Tane
  5. Daisuke Yamashita
  6. Keishi Shintomi
  7. Tatsuya Hirano
(2022)
Molecular dissection of condensin II-mediated chromosome assembly using in vitro assays
eLife 11:e78984.
https://doi.org/10.7554/eLife.78984

Share this article

https://doi.org/10.7554/eLife.78984

Further reading

    1. Chromosomes and Gene Expression
    2. Genetics and Genomics
    Omid Gholamalamdari, Tom van Schaik ... Andrew S Belmont
    Research Article

    Models of nuclear genome organization often propose a binary division into active versus inactive compartments yet typically overlook nuclear bodies. Here, we integrated analysis of sequencing and image-based data to compare genome organization in four human cell types relative to three different nuclear locales: the nuclear lamina, nuclear speckles, and nucleoli. Although gene expression correlates mostly with nuclear speckle proximity, DNA replication timing correlates with proximity to multiple nuclear locales. Speckle attachment regions emerge as DNA replication initiation zones whose replication timing and gene composition vary with their attachment frequency. Most facultative LADs retain a partially repressed state as iLADs, despite their positioning in the nuclear interior. Knock out of two lamina proteins, Lamin A and LBR, causes a shift of H3K9me3-enriched LADs from lamina to nucleolus, and a reciprocal relocation of H3K27me3-enriched partially repressed iLADs from nucleolus to lamina. Thus, these partially repressed iLADs appear to compete with LADs for nuclear lamina attachment with consequences for replication timing. The nuclear organization in adherent cells is polarized with nuclear bodies and genomic regions segregating both radially and relative to the equatorial plane. Together, our results underscore the importance of considering genome organization relative to nuclear locales for a more complete understanding of the spatial and functional organization of the human genome.

    1. Chromosomes and Gene Expression
    Ashwin Govindan, Nicholas K Conrad
    Research Article

    O-GlcNAcylation is the reversible post-translational addition of β-N-acetylglucosamine to serine and threonine residues of nuclear and cytoplasmic proteins. It plays an important role in several cellular processes through the modification of thousands of protein substrates. O-GlcNAcylation in humans is mediated by a single essential enzyme, O-GlcNAc transferase (OGT). OGT, together with the sole O-GlcNAcase OGA, form an intricate feedback loop to maintain O-GlcNAc homeostasis in response to changes in cellular O-GlcNAc using a dynamic mechanism involving nuclear retention of its fourth intron. However, the molecular mechanism of this dynamic regulation remains unclear. Using an O-GlcNAc responsive GFP reporter cell line, we identify SFSWAP, a poorly characterized splicing factor, as a trans-acting factor regulating OGT intron detention. We show that SFSWAP is a global regulator of retained intron splicing and exon skipping that primarily acts as a negative regulator of splicing. In contrast, knockdown of SFSWAP leads to reduced inclusion of a ‘decoy exon’ present in the OGT retained intron which may mediate its role in OGT intron detention. Global analysis of decoy exon inclusion in SFSWAP and UPF1 double knockdown cells indicate altered patterns of decoy exon usage. Together, these data indicate a role for SFSWAP as a global negative regulator of pre-mRNA splicing and positive regulator of intron retention.