Measuring mechanical strains in live cells with force nanosensors

Membrane adhesion receptors and cytoskeleton are two key elements in cell biology mechanotransduction. The molecular machinery of mechanotransduction is a transversal hot topic that give rise to intense activity in scientific theme mixing physics, chemistry , biochemistry and biology . Our ability to understand mechanical cues in biology is limited by our capability to measure it in living cells. Therefore, the spatio-temporal dynamics and regulation of active zones involved in mechanosensing, typically at focal adhesion sites, are still poorly understood. This collaborative project proposes to develop a new force nanosensor (allowing to measure and map the mechanical strains exerted by living cell. This probe is composed of an extensible elastic molecule, flanked by two emitters at its ends, including a quantum dot. The force measurement will be achieved when the elastic molecule is stretched, and Förster Resonant Energy Transfer (FRET) will constitute the way to quantify this stretching at the nanoscale. In this project we also propose to highlight the benefits of our new force nanosensor to probe mechanosensing processes by developing two original applications. These applications will require to implement force nanosensors for super-resolution fluorescence imaging. The first application will use a biomimetic platform allowing to investigate the dynamics of cytoskeleton contractility on integrin focal adhesion complex. The second application will consist in measuring the dynamic of contractile forces in 2D environment, generated by fibroblasts.

Full proposal: https://lightinparis.eu/measuring-mechanical-strains-in-live-cells-with-force

Complementarity between supervisors and interdisciplinarity

Marcelina Cardoso Dos Santois (MCDS) is a cell biologist, while Rodolphe Jaffiol (RF) is a biophysicist. Both partners work in the field of biophysics and have an interdisciplinary background mixing physics, biology, chemistry and biochemistry. They had previously worked together to develop a nano-imaging technique called variable-angle TIRF, allowing to quantify cell binding energy. This new collaborative project is in line with the condition of interdisciplinarity ("3i"). In fact, both partners are associated with two different doctoral schools.

Resources

The project is already supported by an ANR funding (ANR-23-CE42-0009-01). QD-based force sensors are already available. The microscopy techniques such as single molecule fluorescence imaging, FRAP, FRET, TIRF and optical tweezers are already implemented in partner’s labs. Both labs have as well access to cell culture unit. Protein purification will be implemented in MCDS lab.

Supervision experience

Both partners have extensive experience in doctoral supervision and hold Habilitation to Direct Research (HDR) degrees. RJ has supervised 11 doctoral theses. 8 have been defended and 3 are in progress. The MCDS supervised 4 PhDs, 2 of which were completed and 2 of which continue to progress. The partners have indicated their willingness to undertake the supervision of this project, with an allocation of 50% and 30% full-time equivalent (FTE) for MCDS and RJ, respectively.

This interdisciplinary PhD project will mobilise ground-breaking imaging techniques such as :
- single molecule fluorescence microscopy 
- Förster resonance energy transfer (FRET)
- Optical tweezers coupled force spectroscopy
- Fluorescence Recovery After Photobleaching (FRAP)
- Total Internal Reflection Fluorescence Microscopy (TIRFM)
In vitro and in cell experiments:
- Cell culture
- In vitro biomimetic systems (supported lipid bilayers)
Biochemistry:
- Protein purification
Data Analysis:
- Deep learning 

We are therefore looking for a highly motivated, inquisitive candidate who wishes to work in an interdisciplinary environment to complete a PhD at the interface between cell biology, biophysics and biochemistry. The candidate may have an initial background in biology, physics or biophysics with a strong aptitude for experimental work.