Mapping molecular assemblies with fluorescence microscopy and object-based spatial statistics


Thibault Lagache 1,5, Alexandre Grassart2, Stéphane Dallongeville1, Orestis Faklaris3, Nathalie Sauvonnet2,
Alexandre Dufour1, Lydia Danglot 4 & Jean-Christophe Olivo-Marin 1

1 Institut Pasteur, BioImage Analysis Unit. CNRS UMR 3691. 25 rue du Docteur Roux, 75724 Paris Cedex 15, France.
2 Institut Pasteur, Molecular Microbial Pathogenesis Unit. INSERM U1202. 28 rue du Docteur Roux, 75724 Paris Cedex 15, France.
3 CNRS UMR7592, Institut Jacques Monod, Université Paris Diderot, 15 rue Hélène Brion, 75013 Paris, France.
4 Inserm U894 Center for Psychiatry and Neuroscience, Team Membrane traffic in healthy and diseased
brain, 102-108 rue de la Santé, 75014 Paris, France.
5 Present address: Department of Biological Sciences, Columbia University, New York NY USA.


Elucidating protein functions and molecular organisation requires to localise precisely single or aggregated molecules and analyse their spatial distributions. We develop a statistical method SODA (Statistical Object Distance Analysis) that uses either micro- or nanoscopy to significantly improve on standard co-localisation techniques. Our method considers cellular geometry and densities of molecules to provide statistical maps of isolated and associated (coupled) molecules. We use SODA with three-colour structured-illumination microscopy (SIM) images of hippocampal neurons, and statistically characterise spatial organisation of thousands of synapses. We show that presynaptic synapsin is arranged in asymmetric triangle with the 2 postsynaptic markers homer and PSD95, indicating a deeper localisation of homer. We then determine stoichiometry and distance between localisations of two synaptic vesicle proteins with 3D-STORM. These findings give insights into the protein organisation at the synapse, and prove the efficiency of SODA to quantitatively assess the geometry of molecular assemblies.

NATURE COMMUNICATIONS | (2018) 9:698 | DOI: 10.1038/s41467-018-03053-x |

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