1. Cell Biology

Splicing factors Sf3A2 and Prp31 have direct roles in mitotic chromosome segregation

  1. Claudia Pellacani
  2. Elisabetta Bucciarelli
  3. Fioranna Renda
  4. Daniel Hayward
  5. Antonella Palena
  6. Jack Chen
  7. Silvia Bonaccorsi
  8. James G Wakefield
  9. Maurizio Gatti  Is a corresponding author
  10. Maria Patrizia Somma  Is a corresponding author
  1. University of Rome, Italy
  2. University of Exeter, United Kingdom
https://doi.org/10.7554/eLife.40325
Share this article
Cite this article
  1. Claudia Pellacani
  2. Elisabetta Bucciarelli
  3. Fioranna Renda
  4. Daniel Hayward
  5. Antonella Palena
  6. Jack Chen
  7. Silvia Bonaccorsi
  8. James G Wakefield
  9. Maurizio Gatti
  10. Maria Patrizia Somma
(2018)
Splicing factors Sf3A2 and Prp31 have direct roles in mitotic chromosome segregation
eLife 7:e40325.
https://doi.org/10.7554/eLife.40325
  2. Download BibTeX
  3. Download .RIS

Abstract

Several studies have shown that RNAi-mediated depletion of splicing factors (SFs) results in mitotic abnormalities. However, it is currently unclear whether these abnormalities reflect defective splicing of specific pre-mRNAs or a direct role of the SFs in mitosis. Here we show that two highly conserved SFs, Sf3A2 and Prp31, are required for chromosome segregation in both Drosophila and human cells. Injections of anti-Sf3A2 and anti-Prp31 antibodies into Drosophila embryos disrupt mitotic division within 1 minute, arguing strongly against a splicing-related mitotic function of these factors. We demonstrate that both SFs bind spindle microtubules (MTs) and the Ndc80 complex, which in Sf3A2- and Prp31-depleted cells is not tightly associated with the kinetochores; in HeLa cells the Ndc80/HEC1-SF interaction is restricted to the M phase. These results indicate that Sf3A2 and Prp31 directly regulate interactions among kinetochores, spindle microtubules and the Ndc80 complex in both Drosophila and human cells.

Data availability

All data generated or analyzed during this study are included in the manuscript and Supplementary File 1. The full dataset for proteomic analyses reported in Figures 6 and Supplementary File 1 can be found at https://www.thewakefieldlab.com/ms; the significantly reduced protein IDs for each RNAi experiment were interrogated using a Gene Ontology (GO) classifier (GOTermMapper (https://go.princeton.edu/cgi-bin/GOTermMapper), concentrating on the GO terms ""cell division"" (GO:0051301), ""mitotic cell cycle"" (GO:0000278) and ""chromosome segregation"" (GO:0007059). Source data files have been provided for Figures 1, 4, 5, 7, 8, and 9.

Article and author information

Author details

  1. Claudia Pellacani

    Department of Biology and Biotechnology Charles Darwin, University of Rome, Rome, Italy
    Competing interests
    The authors declare that no competing interests exist.

  2. Elisabetta Bucciarelli

    Institute of Molecular Biology and Pathology, University of Rome, Rome, Italy
    Competing interests
    The authors declare that no competing interests exist.

  3. Fioranna Renda

    Department of Biology and Biotechnology Charles Darwin, University of Rome, Rome, Italy
    Competing interests
    The authors declare that no competing interests exist.

  4. Daniel Hayward

    Biosciences/Living Systems Institute, University of Exeter, Exeter, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.

  5. Antonella Palena

    Institute of Molecular Biology and Pathology, University of Rome, Rome, Italy
    Competing interests
    The authors declare that no competing interests exist.

  6. Jack Chen

    Biosciences/Living Systems Institute, University of Exeter, Exeter, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.

  7. Silvia Bonaccorsi

    Department of Biology and Biotechnology Charles Darwin, University of Rome, Roma, Italy
    Competing interests
    The authors declare that no competing interests exist.

  8. James G Wakefield

    Biosciences/Living Systems Institute, University of Exeter, Exeter, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.

  9. Maurizio Gatti

    Department of Biology and Biotechnology Charles Darwin, University of Rome, Rome, Italy
    For correspondence
    maurizio.gatti@uniroma1.it
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-3777-300X

  10. Maria Patrizia Somma

    Institute of Molecular Biology and Pathology, University of Rome, Rome, Italy
    For correspondence
    patrizia.somma@uniroma1.it
    Competing interests
    The authors declare that no competing interests exist.

Funding

Associazione Italiana per la Ricerca sul Cancro (IG16020)

  • Maurizio Gatti

Ministero dell'Istruzione, dell'Università e della Ricerca

  • Silvia Bonaccorsi

Biotechnology and Biological Sciences Research Council (BB/K017837/1)

  • James G Wakefield

Associazione Italiana per la Ricerca sul Cancro (IG20528)

  • Maurizio Gatti

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

© 2018, Pellacani 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

  • 2,315
    views
  • 348
    downloads
  • 20
    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. Claudia Pellacani
  2. Elisabetta Bucciarelli
  3. Fioranna Renda
  4. Daniel Hayward
  5. Antonella Palena
  6. Jack Chen
  7. Silvia Bonaccorsi
  8. James G Wakefield
  9. Maurizio Gatti
  10. Maria Patrizia Somma
(2018)
Splicing factors Sf3A2 and Prp31 have direct roles in mitotic chromosome segregation
eLife 7:e40325.
https://doi.org/10.7554/eLife.40325

Share this article

https://doi.org/10.7554/eLife.40325

Categories and tags

Research organism

Further reading

    1. Cell Biology
    2. Neuroscience

    A neurotrophin functioning with a Toll regulates structural plasticity in a dopaminergic circuit

    Jun Sun, Francisca Rojo-Cortes ... Alicia Hidalgo
    Research Article

    Experience shapes the brain as neural circuits can be modified by neural stimulation or the lack of it. The molecular mechanisms underlying structural circuit plasticity and how plasticity modifies behaviour are poorly understood. Subjective experience requires dopamine, a neuromodulator that assigns a value to stimuli, and it also controls behaviour, including locomotion, learning, and memory. In Drosophila, Toll receptors are ideally placed to translate experience into structural brain change. Toll-6 is expressed in dopaminergic neurons (DANs), raising the intriguing possibility that Toll-6 could regulate structural plasticity in dopaminergic circuits. Drosophila neurotrophin-2 (DNT-2) is the ligand for Toll-6 and Kek-6, but whether it is required for circuit structural plasticity was unknown. Here, we show that DNT-2-expressing neurons connect with DANs, and they modulate each other. Loss of function for DNT-2 or its receptors Toll-6 and kinase-less Trk-like kek-6 caused DAN and synapse loss, impaired dendrite growth and connectivity, decreased synaptic sites, and caused locomotion deficits. In contrast, over-expressed DNT-2 increased DAN cell number, dendrite complexity, and promoted synaptogenesis. Neuronal activity modified DNT-2, increased synaptogenesis in DNT-2-positive neurons and DANs, and over-expression of DNT-2 did too. Altering the levels of DNT-2 or Toll-6 also modified dopamine-dependent behaviours, including locomotion and long-term memory. To conclude, a feedback loop involving dopamine and DNT-2 highlighted the circuits engaged, and DNT-2 with Toll-6 and Kek-6 induced structural plasticity in this circuit modifying brain function and behaviour.

    1. Cell Biology

    Inclusive, exclusive and hierarchical atlas of NFATc1+/PDGFR-α+ cells in dental and periodontal mesenchyme

    Xue Yang, Chuyi Han ... Fanyuan Yu
    Research Article

    Platelet-derived growth factor receptor alpha (PDGFR-α) activity is crucial in the process of dental and periodontal mesenchyme regeneration facilitated by autologous platelet concentrates (APCs), such as platelet-rich fibrin (PRF), platelet-rich plasma (PRP) and concentrated growth factors (CGF), as well as by recombinant PDGF drugs. However, it is largely unclear about the physiological patterns and cellular fate determinations of PDGFR-α+ cells in the homeostasis maintaining of adult dental and periodontal mesenchyme. We previously identified NFATc1 expressing PDGFR-α+ cells as a subtype of skeletal stem cells (SSCs) in limb bone in mice, but their roles in dental and periodontal remain unexplored. To this end, in the present study we investigated the spatiotemporal atlas of NFATc1+ and PDGFR-α+ cells residing in dental and periodontal mesenchyme in mice, their capacity for progeny cell generation, and their inclusive, exclusive and hierarchical relations in homeostasis. We utilized CRISPR/Cas9-mediated gene editing to generate two dual recombination systems, which were Cre-loxP and Dre-rox combined intersectional and exclusive reporters respectively, to concurrently demonstrate the inclusive, exclusive, and hierarchical distributions of NFATc1+ and PDGFR-α+ cells and their lineage commitment. By employing the state-of-the-art transgenic lineage tracing techniques in cooperating with tissue clearing-based advanced imaging and three-dimensional slices reconstruction, we systematically mapped the distribution atlas of NFATc1+ and PDGFR-α+ cells in dental and periodontal mesenchyme and tracked their in vivo fate trajectories in mice. Our findings extend current understanding of NFATc1+ and PDGFR-α+ cells in dental and periodontal mesenchyme homeostasis, and furthermore enhance our comprehension of their sustained therapeutic impact for future clinical investigations.

    1. Cell Biology
    2. Neuroscience

    Endosomal sorting protein SNX4 limits synaptic vesicle docking and release

    Josse Poppinga, Nolan J Barrett ... Jan RT van Weering
    Research Article

    Sorting nexin 4 (SNX4) is an evolutionary conserved organizer of membrane recycling. In neurons, SNX4 accumulates in synapses, but how SNX4 affects synapse function remains unknown. We generated a conditional SNX4 knock-out mouse model and report that SNX4 cKO synapses show enhanced neurotransmission during train stimulation, while the first evoked EPSC was normal. SNX4 depletion did not affect vesicle recycling, basic autophagic flux, or the levels and localization of SNARE-protein VAMP2/synaptobrevin-2. However, SNX4 depletion affected synapse ultrastructure: an increase in docked synaptic vesicles at the active zone, while the overall vesicle number was normal, and a decreased active zone length. These effects together lead to a substantially increased density of docked vesicles per release site. In conclusion, SNX4 is a negative regulator of synaptic vesicle docking and release. These findings suggest a role for SNX4 in synaptic vesicle recruitment at the active zone.