Alpha-Synuclein nucleates short microtubules

Alpha-Synuclein (alpha-syn) is the major component of Lewy body, the cytosolic inclusions associated to Parkinson’s disease, and its mutation have been linked to genetic forms of the disorder. Although largely studied, alpha-syn physiological role is still unclear. In vitro, -syn is unfolded, but it acquires a secondary structure upon interaction with the partners. It is also reported the capacity of alpha-syn to induce microtubule (MT) polymerization, but a detailed analysis of kinetic parameters still lacks. Here, we decided to undertake an in depth evaluation of the effects of alpha-syn on tubulin polymerization kinetics. Because standard protocol failed in reproducibility, we decided to incubate alpha-syn in the presence of tubulin, just to allow alpha-syn folding. Circular dichroism spectra show that upon pre-incubation alpha-syn acquires a -helix conformation. Thus, we performed tubulin polymerization following pre-incubation with WT and mutated alpha-syns, and the analyses of kinetic parameters suggest that synucleins promote MT nucleation, but they seem to reduce MT mass at the end of polymerization. By using fluorescence microscopy, we evaluated the number and the length of MTs polymerized in presence of alpha-syns, at the very beginning and at the end of the polymerization. Accordingly to the already published data, mutated alpha-syn causes tubulin aggregation instead of MT polymerization. On the contrary, at both time points, WT alpha-syn promotes the polymerization of a higher number of MTs, confirming its capacity in nucleating MTs. Moreover, MTs polymerized in the presence of WT alpha-syn are shorter in respect to control MTs, reconciling the apparent reduction of MT mass due to the physics of the system. Our results underscore the ability of alpha-syn to nucleate short MTs, as usually are MTs present at synaptic terminal, the cell compartment where alpha-syn resides. Thus, we shed light on a new physiological role for alpha-syn as a feasible axonal MT nucleating structures.