A robust and tunable mitotic oscillator in artificial cells

  1. Ye Guan
  2. Zhengda Li
  3. Shiyuan Wang
  4. Patrick M Barnes
  5. Xuwen Liu
  6. Haotian Xu
  7. Minjun Jin
  8. Allen P Liu
  9. Qiong Yang  Is a corresponding author
  1. University of Michigan, United States
  2. University of Science and Technology of China, China
  3. Wayne State University, United States

Abstract

Single-cell analysis is pivotal to deciphering complex phenomena like heterogeneity, bistability, and asynchronous oscillations, where a population ensemble cannot represent individual behaviors. Bulk cell-free systems, despite having unique advantages of manipulation and characterization of biochemical networks, lack the essential single-cell information to understand a class of out-of-steady-state dynamics including cell cycles. Here, by encapsulating Xenopus egg extracts in water-in-oil microemulsions, we developed artificial cells that are adjustable in sizes and periods, sustain mitotic oscillations for over 30 cycles, and function in forms from the simplest cytoplasmic-only to the more complicated ones involving nuclear dynamics, mimicking real cells. Such innate flexibility and robustness make it key to studying clock properties like tunability and stochasticity. Our results also highlight energy as an important regulator of cell cycles. We demonstrate a simple, powerful, and likely generalizable strategy of integrating strengths of single-cell approaches into conventional in vitro systems to study complex clock functions.

Data availability

The following data sets were generated

Article and author information

Author details

  1. Ye Guan

    Department of Biophysics, University of Michigan, Ann Arbor, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Zhengda Li

    Department of Biophysics, University of Michigan, Ann Arbor, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Shiyuan Wang

    Department of Biophysics, University of Michigan, Ann Arbor, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Patrick M Barnes

    Department of Physics, University of Michigan, Ann Arbor, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Xuwen Liu

    Department of Physics, University of Science and Technology of China, Hefei, China
    Competing interests
    The authors declare that no competing interests exist.
  6. Haotian Xu

    Department of Computer Science, Wayne State University, Detroit, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. Minjun Jin

    Department of Biological Chemistry, University of Michigan, Ann Arbor, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Allen P Liu

    Department of Biophysics, University of Michigan, Ann Arbor, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0309-7018
  9. Qiong Yang

    Department of Biophysics, University of Michigan, Ann Arbor, United States
    For correspondence
    qiongy@umich.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2442-2094

Funding

National Institutes of Health (MIRA #GM119688)

  • Qiong Yang

National Science Foundation (CAREER Grant #1553031)

  • Qiong Yang

Alfred P. Sloan Foundation (Sloan Research Fellowship)

  • Qiong Yang

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

Copyright

© 2018, Guan 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

  • 4,274
    views
  • 559
    downloads
  • 42
    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. Ye Guan
  2. Zhengda Li
  3. Shiyuan Wang
  4. Patrick M Barnes
  5. Xuwen Liu
  6. Haotian Xu
  7. Minjun Jin
  8. Allen P Liu
  9. Qiong Yang
(2018)
A robust and tunable mitotic oscillator in artificial cells
eLife 7:e33549.
https://doi.org/10.7554/eLife.33549

Share this article

https://doi.org/10.7554/eLife.33549

Further reading

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics
    Assmaa Elsheikh, Camden M Driggers ... Show-Ling Shyng
    Research Article

    Pancreatic KATP channel trafficking defects underlie congenital hyperinsulinism (CHI) cases unresponsive to the KATP channel opener diazoxide, the mainstay medical therapy for CHI. Current clinically used KATP channel inhibitors have been shown to act as pharmacochaperones and restore surface expression of trafficking mutants; however, their therapeutic utility for KATP trafficking-impaired CHI is hindered by high affinity binding, which limits functional recovery of rescued channels. Recent structural studies of KATP channels employing cryo-electron microscopy (cryoEM) have revealed a promiscuous pocket where several known KATP pharmacochaperones bind. The structural knowledge provides a framework for discovering KATP channel pharmacochaperones with desired reversible inhibitory effects to permit functional recovery of rescued channels. Using an AI-based virtual screening technology AtomNet followed by functional validation, we identified a novel compound, termed Aekatperone, which exhibits chaperoning effects on KATP channel trafficking mutations. Aekatperone reversibly inhibits KATP channel activity with a half-maximal inhibitory concentration (IC50) ~9 μM. Mutant channels rescued to the cell surface by Aekatperone showed functional recovery upon washout of the compound. CryoEM structure of KATP bound to Aekatperone revealed distinct binding features compared to known high affinity inhibitor pharmacochaperones. Our findings unveil a KATP pharmacochaperone enabling functional recovery of rescued channels as a promising therapeutic for CHI caused by KATP trafficking defects.

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics
    Andrew P Latham, Longchen Zhu ... Bin Zhang
    Research Article

    The phase separation of intrinsically disordered proteins is emerging as an important mechanism for cellular organization. However, efforts to connect protein sequences to the physical properties of condensates, that is, the molecular grammar, are hampered by a lack of effective approaches for probing high-resolution structural details. Using a combination of multiscale simulations and fluorescence lifetime imaging microscopy experiments, we systematically explored a series of systems consisting of diblock elastin-like polypeptides (ELPs). The simulations succeeded in reproducing the variation of condensate stability upon amino acid substitution and revealed different microenvironments within a single condensate, which we verified with environmentally sensitive fluorophores. The interspersion of hydrophilic and hydrophobic residues and a lack of secondary structure formation result in an interfacial environment, which explains both the strong correlation between ELP condensate stability and interfacial hydrophobicity scales, as well as the prevalence of protein-water hydrogen bonds. Our study uncovers new mechanisms for condensate stability and organization that may be broadly applicable.