High-speed nanoscopy to decipher the real-time mitochondrial dynamics - Dyn@mit

Contact: Stéphanie Bonneau

Dyn@mit team -

The research activities of the Dyn@mit project-team are conducted within UMRs, under the joint supervision of UPMC and CNRS. The two laboratories involved have activities in physics and imaging. The LPEM presents a set of recognized skills and technical know-how, for example by mastering advanced experimental imaging methods. Within the Dyn@mit project, the latter meet the activities of the LJP, which come within physics at the interface with biology and aim at exploring the response of a biological systems to external stresses and perturbations. In this line, the Dyn@mit approaches allow to probe the properties of a complex biological system, the mitochondria, at different scales, to give new insights on its functioning.

Dyn@mit project -

Mitochondria are key subcellular structures in eukaryote cells. First, they are considered as the cell's powerhouse, whose particular architecture and biochemical properties serve one major purpose, the  optimization of energy production by oxidative phosphorylation. Moreover, it became clear during the 1990's that mitochondria have a second crucial function as they are the regulation center of cell death [Kroemer 2007]. If this idea at first appeared counterintuitive (How could the mitochondrial vital forces be turn away to serve a lethal purpose),  the link between the two mitochondrial functions is today viewed as a major key of cell's regulation. Indeed, in healthy cells, the integrity of mitochondrial membranes ensures the establishment of transmembrane gradients subsequently exploited by the respiratory chain to drive energy. Their resistance to unusual conditions guarantees the robustness of living cells. Conversely, when the stress becomes too important to preserve integrity and functionality of these structures, their destabilization is the first cell death signal that enable the disruption of pathological cells - what may preserve the organism survival. Understanding the interrelationships between these structures and the mitochondrial functions is then a major issue for cellular biologists and biophysicists.

Electron microscopy imaging led, from the 1950's to the 1990's, to the canonical representation of mitochondria as bean-shaped organelles of which hallmarks are the double membranes and unusual inner membrane nano-sized involutions, so-called cristae. Recent years have renewed our vision of the remarkably dynamic nature of mitochondria. Theses organelles constantly change shape, fuse and divide, and are remodeled even in their inner nanostructure [Detmer 2007]. Since the late 2000's, electron tomography studies strongly suggest that inner membrane topology is a regulated property of mitochondria. Arguments were given to support the idea that physiological factors determine this membrane's topology and, conversely, that the inner membrane shape influences mitochondrial functions. Such dynamics is crucial for eukaryotic living organisms, and defects lead to important disorders (neurodegenerative diseases, cancer and ageing, for example). But what are the molecular processes controlling this dynamics, and why is the latter essential for mitochondrial function ?

Scientific objectives -

Therefore, mitochondrial functions seems closely dependent on a particular multi-scale topology. Although mechanisms that regulate mitochondrial abundance and shape are qualitatively understood, little is known about how mitochondrial function and nanostructure are inter-linked and regulated. In particular, the dynamics of mitochondrial inner compartments remodeling is still not known because unattainable by conventional imaging techniques. Our goal is to overcome these technical limits to decipher real-time mitochondrial dynamics at the nano-scale and to give new insights into the functional and regulator role of their nanostructure.

Leading-edge imaging technique -

Fluorescence microscopy is now the imaging method of choice in biology. It has several advantages: sensitivity, flexibility, sufficient acquisition speed to study the remodeling of biological systems, in space and in time. It suffers however from a limit of resolution inherent to conventional optical systems. Due to the size of mitochondria and of their ultrastructure (cristae), their topological study is impossible by such methods. Nevertheless, the resolution limit can be extended by a structured illumination technique (so-called SIM, "structured-illumination microscopy") which makes possible to encode the high frequencies of the object in the wide field image. Several images of the same sample are acquired for different illumination patterns and are digitally combined to reconstruct a super-resolution image of the object. The resolution can be improved by a factor of two. If seven images are typically needed, we developed a reconstruction method based on only four images, improving the temporal resolution of the acquisition, essential for the proposed study.

Mitochondrial remodeling -

We can control the cells physiological state by using a photoactivable molecule, which activation allows to tune the level of cell's stress [Aubertin 2013]. Such molecules were previously used to photo-control the inner cell trafficking [Sultan 2016].

Links -


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