Interaction neurones-microglies via les Tunneling Nanotubes (TNTs) en cas de stress synucléinopathique

Intercellular communication is of critical importance in maintenance of homeostasis. Our group and others have identified Tunneling Nanotubes (TNTs) to play significant roles in mediating communication between different cells of the brain such as neurons, astrocytes, and microglia. TNTs are long, actin-rich and membranous intercellular connections. Functionally different from other cellular protrusions, TNTs facilitate material exchange between connected cell - Ca2+ ions, RNAs, viruses, bacteria, organelles (lysosomes and mitochondria), and native proteins and aggregated forms. This defining function of TNTs positions them as an important regulator of health and disease. Neurodegeneration-associated toxic protein aggregates such as in Prion's (PrPSc), Alzheimer's (Amyloid-β and Tau), Parkinson's (PD; α-Synuclein), and Huntington's (mHTT) spread through TNTs. α-Synuclein (α-Syn), for example, hijack TNTs to propagate between neurons, astrocytes, microglia, and from neurons to astrocytes. However, the role of TNTs in mediating neuron-microglia interaction had remained elusive, forming the basis of this work.Using human cell lines SH-SY5Y (neuronal) and HMC3 (microglial), I initially demonstrated functional TNTs between the two cell types in a PD-mimicking condition involving exogenous pre-formed fibrils of α-Syn. These aggregates increased, and hijacked TNTs for propagation between cells. Neuronal cells were efficient in transferring α-Syn to microglia, while microglia transferred mitochondria preferably to α-Syn burdened neuronal cells. Next, I demonstrated that directional trafficking of α-Syn from neuronal cells to microglia was a consequence of lysosomal damage and autophagy dysfunction. In contrast, α-Syn-exposed microglia had limited compromise of lysosomal degradative ability, exhibited efficient lysosomal turnover and autophagy flux. Similar phenotype was observed in iPSC-derived neurons and microglia. Microglia also upregulated autophagy when co-cultured with α-Syn-laden neuronal cells, that enabled efficient degradation of transferred aggregates. α-Syn aggregates also caused significant damage to mitochondria, compromising membrane integrity of these organelles that led to release of damage associated molecular patterns such as mitochondrial DNA in the cytosol. I showed that this contributed to the activation of the innate immune pathway cGAS-STING, both in cell lines and iPSC-derived cells, which positively regulated the formation of intercellular connections. Downstream activation of the major inflammatory transcription factor NF-κB, however, represents the key step in the process of formation of TNTs. This not only resulted in higher aggregate spread, but also of damaged mitochondria from neuronal cells to microglia, which were eventually targeted for lysosome-mediated clearance. My dissertation work lead to a postulate that TNTs represent a ‘dual-traffic highway' in the context of neuron-microglia interactions - allowing neuronal cells to dissipate accumulated aggregates and damaged organelles, while enabling potential trophic support from microglia to aggregate-burdened neuronal cells both directly (by providing functional mitochondria) and indirectly (by degrading neuronal cell-derived α-Syn and damaged mitochondria).