α-Synuclein interacts differently with membranes mimicking the inner and outer leaflets of neuronal membranes.

The toxicity of α-synuclein (α-syn), the amyloidogenic protein responsible for Parkinson's disease, is likely related to its interaction with the asymmetric neuronal membrane. α-Syn exists as cytoplasmatic and as extracellular protein as well. To shed light on the different interactions occurring at the different α-syn localizations, we have here modelled the external and internal membrane leaflets of the neuronal membrane with two complex lipid mixtures, characterized by phase coexistence and with negative charge confined to either the ordered or the disordered phase, respectively. To this purpose, we selected a five-component (DOPC/SM/DOPE/DOPS/chol) and a four-component (DOPC/SM/GM1/chol) lipid mixtures, which contained the main membrane lipid constituents and exhibited a phase separation with formation of ordered domains. We have compared the action of α-syn in monomeric form and at different concentrations (1 nM, 40 nM, and 200 nM) with respect to lipid systems with different composition and shape by AFM, QCM-D, and vesicle leakage experiments. The experiments coherently showed a higher stability of the membranes composed by the internal leaflet mixture to the interaction with α-syn. Damage to membranes made of the external leaflet mixture was detected in a concentration-dependent manner. Interestingly, the membrane damage was related to the fluidity of the lipid domains and not to the presence of negatively charged lipids.

Correlative nanoscopy: A multimodal approach to molecular resolution.

Atomic force microscopy (AFM) is a nano-mechanical tool uniquely suited for biological studies at the molecular scale. AFM operation is based on mechanical interaction between the tip and the sample, a mechanism of contrast capable of measuring different information, including surface topography, mechanical, and electrical properties. However, the lack of specificity highlights the need to integrate AFM data with other techniques providing compositional hints. In particular, optical microscopes based on fluorescence as a mechanism of contrast can access the local distribution of specific molecular species. The coupling between AFM and super-resolved fluorescence microscopy solves the resolution mismatch between AFM and conventional fluorescence optical microscopy. Recent advances showed that also the inherently label-free imaging capabilities of the AFM are fundamental to complement the fluorescence images. In this review, we have presented a brief historical view on correlative microscopy, and, finally, we have summarized the progress of correlative AFM-super-resolution microscopy in biological research.