alpha-synuclein oligomerization in HeLa cells: role of mitochondria on aggregation?

S. GALLI; M. J. ROBERTI; J. H. MARTINEZ; L. I. PIETRASANTA; T. M. JOVIN; E. A. JARES-ERIJMAN

Buzios, Brasil

Congreso; I Congresso IBRO/LARC de Neurociencias da America Latina, Caribe e Península Ibérica.; 2008

Sociedade Brasileira de Neurociencias e Comportamento - SBNeC

alpha-synuclein (a-synuclein) belongs to a small group of natively unfolded proteins that can transiently bind to lipid membranes and acquire a partial a-helical conformation. Under certain pathogenic conditions, a-synuclein associates to form oligomers and insoluble fibrils with increased beta-sheet configuration. Nitration of a-synuclein mediated by ONOO- may play a mechanistic role in PD pathogenesis, i.e. i) nitration induces the formation of a-synuclein oligomers, ii) low concentrations of nitrated protein facilitate the nucleation step of a-synuclein, but iii) at high concentrations, these species inhibit filament assembly, iv) Tyr39 nitration displaces a-synuclein from lipid vesicles, and v) nitrated a-synuclein exhibits a reduced proteosomal degradation. a-synuclein was found in the mitochondria of different cells, and there it regulated mitochondrial NO production, and thus free radical formation. Some authors found that oligomeric a-synuclein bound to mitochondria, but not the monomer. This raises the question of whether mitochondria are capable of accelerating a-synuclein oligomerization, or if a-synuclein binds mitochondria as a pre-formed oligomer. Here we showed that overexpressed a-synuclein entered the mitochondria of HeLa cells, though the endogenous protein was not present in the organelle, but was rather distributed in membrane and nuclei. In addition, by confocal microscopy we observed colocalization of the oligomeric form with mitochondria, but not of the monomer upon overexpression. We have previously engineered a recombinant a-synuclein with a 12 amino acids tag that encodes a binding site for a fluorescent dye, FlASH. With this construct we were able to map the protein to the mitochondria by flow cytometry on the isolated organelle. In future studies we attempt to elucidate whether the mitochondria induces a-synuclein oligomerization or if it is the oligomerized protein that enters the organelle and induces oxidative stress. We also want to elucidate the role of mitochondrial radicals in a-synuclein aggregation. These findings altogether may support a pivotal role for mitochondria and a-synuclein in oxidative stress and apoptosis.