SPECIAL SEMINAR

“Super-resolution Microscopy for the
Direct Visualization of Seeding and
Toxicity of α-Synuclein Species in Neurons”

Friday June 16th 2017
13:00-14:00
BRFAA Amphitheater

Speaker:

Dorothea Pinotsi

ETH Zurich
dpinotsi@ethz.ch

HOST: Lukas H. Margaritis
Professor Emeritus Cell Biology and Electron Microscopy.
Dept. of Biology, Athens University
Affiliated Investigator. BRFAA

For more information, please contact
Dimitris Thanos, Professor-Academician at thanos@bioacademy.gr
and for directions, please visit http://www.bioacademy.gr/contact
 

Abstract:

Visualizing molecular self-assembly has emerged as one of the great challenges in modern biophysics, related to phenomena ranging from nanotechnology applications and bio inspired material science to the formation of molecular machineries in living cells. In particular, an example of biomolecular self-assembly that gives rise to an important class of macromolecular structures is the formation of protein aggregates the so-called amyloid fibrils. Super-resolution microscopy has emerged as a powerful and non-invasive tool for the study of such processes both in vitro, but also as they occur in live cells. In this talk I will discuss the main principles of the super resolution microscopy technique based on single molecule localization. I will then present the application of one such technique, the direct stochastic optical reconstruction microscopy (dSTORM), to determine the morphology of amyloid proteins as it evolves over time in vitro, but also as it emerges in a complex cellular environment. Using two-colour dSTORM we have shown that there is heterogeneity in the growth rates of individual amyloid fibrils which can be attributed to structural polymorphism. In neuronal cells, we have shown that exogenously added α-synuclein seed fibrils primarily elongate by the endogenous α-synuclein, naturally present in neurons. In contrast, exogenously added monomeric α-synuclein induces nucleation of the endogenous protein and leads to apoptosis. The latter is rescued by the addition of seed fibrils, suggesting a neuroprotective role of fibrillar species. The visualization of these effects at the nanoscale opens up new avenues for understanding the links between α-synuclein aggregation and neuronal toxicity.

Key Publications:

[1] D. Pinotsi, G. S. Kaminski Schierle and C. F. Kaminski, Optical Super-Resolution Imaging of β- Amyloid Aggregation In Vitro and In Vivo: Method and Techniques, Book chapter in Methods Mol Biol.;1303:125-41 (2015).

[2] D. Pinotsi, A. K. Buell, C. Galvagnion, C. M. Dobson, G. S. Kaminski Schierle, and C. F. Kaminski, ''Direct observation of heterogeneous amyloid fibril growth kinetics via two-color super-resolution microscopy'', Nano Letters 14, 339-345 (2014).

[3] D. Pinotsi, L. Grisanti, P. Mahou, R. Gebauer, C. F. Kaminski, A. Hassanali, G. S. Kaminski Schierle, ''Proton transfer and structure-specific fluorescence in hydrogen bond–rich protein structures'', Journal of the American Chemical Society (JACS) 138 (9), pp 3046–3057 (2016).

[4] D. Pinotsi, C. H. Michel, A. K. Buell, R.F. Laine, P. Mahou, C. M. Dobson, C. F. Kaminski, G. S. Kaminski Schierle, ''Nanoscopic insights into seeding mechanisms and toxicity of alpha-synuclein species in neurons'', Proceedings of the National Academy of Sciences of the United States of America, (PNAS) 113 (14), pp 3815–3819 (2016).

[5] L. Grisanti, D. Pinotsi, R. Gebauer, G.S. Kaminski Schierle, A. Hassanali, ‘’A computational study on how structure influences the optical properties in model crystal structures of amyloid fibrils.’’
Phys Chem Chem Phys, 19(5):4030-4040. doi: 10.1039/c6cp07564a, (2017).