Collaborative Research: Uncovering the Biophysical Mechanisms of Single-cell Wound-healing

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Creating systems that confer life-like properties has been a major endeavor in both science and engineering. Recent years have witnessed an explosion in research on building synthetic cells, not only as a way to study the origins and the rules of life, but also as a new approach to biochemical engineering to increase the yield in making molecules. Nevertheless, current synthetic cell research has neglected one of the most fundamental properties of living matter – the ability to self-repair following damage. Living cells are generally soft and easily damaged, yet a number of them can repair themselves after being mechanically punctured, torn, or even ripped in half. If one could construct such self-repairing capability in synthetic cells, it should be possible to gain insights into one of the defining features of cells. At the same time, it could open new realms of biochemical engineering by allowing the synthetic cell systems to operate robustly under the potentially harsh environment of industrial processes. One approach to attaining self-repairing synthetic cells is to adapt self-repair mechanisms from a living system which already demonstrates robust capacity to heal from large mechanical wounds within a single cell, and build analogs of these mechanisms inside synthetic cells. One such system is Stentor coeruleus, a single-celled free-living ciliate, which possess a more robust wound healing capacity than most other cells. However, the self-repair mechanisms of Stentor are largely unknown. The overall goal of this proposal, therefore, is to understand the mechanisms by which Stentor cells can heal robustly from large mechanical wounds. The results of this work will lead to fundamental insights into wound healing, one of the defining features of life. It will also lay the foundation for constructing self-repairing synthetic cells. The collaboration between the researchers will provide a unique opportunity for training and workforce development at the interface of cell biology and engineering. A website on “superhero cells and bugs” will be created to raise public interest in non-model organisms possessing “superpowers” such as self-healing and survival in space. The researchers will continue their efforts to recruit underrepresented minorities to science and engineering via social media, outreach activities targeted to K-12 students, and their active participation in the Bay Area Science Festival and the Maker Faire.The overall goal of this research is to understand at a physical and molecular level how Stentor coeruleus cells can heal robustly from large mechanical wounds that cause an opening in the plasma membrane. The key biological questions probed include: What sets the limit of the biggest wound the cell can recover from? Does the large size of the cell facilitate its wound healing, or has wound healing evolved to be particularly rapid in this cell? The rationale to focus on Stentor are: 1) its wound healing capacity is more robust than most other cells, capable of recovering from drastic wounds and regenerating from cell fragments as small as 1/27th of original cell size in 24 hours. 2) The ability to perform high-throughput gene knockdown and wounding experiments. The research objectives are to: 1) Develop a minimalistic whole-cell mathematical model of single-cell wound healing. 2) Test predictions of the model by measuring the kinetics of healing in cells as a function of wound size and cell size. 3) Identify contributions to the healing process from membrane patching, purse-string constriction, or other mechanisms as identified by phosphoproteomics. The intellectual merit of this research lies in the identification of the principles for repairing large mechanical wounds in a single cell, and the conditions at which the healing process will succeed or fail. Fundamentally, the ability to heal is one of the key features that distinguish living matter from non-living matter. This study will shed light into the problem of how some biological systems can heal more robustly than others. Practically, the work will lay the foundation for engineering a new function—self-repair—in synthetic cells, and will make the technology more robust for potential scale-up for practical industrial applications.This award was co-funded by the Cellular Dynamics and Function and Systems and Synthetic Biology clusters of the Division of Molecular and Cellular Biosciences.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.