Traffic jams and the complex role of α-Synuclein aggregation in Parkinson disease

Abstract

A common pathological event among various neurodegenerative disorders (NDs) is the misfolding and aggregation of different proteins in the brain. This is thought to potentiate aberrant protein-protein interactions that culminate in the disruption of several biological processes and, ultimately, in neuronal cell loss. Although protein aggregates are a common hallmark in several disorders, the molecular pathways leading to their generation remain unclear. The misfolding and aggregation of α-Synuclein (aSyn) is the pathological hallmark of Parkinson disease (PD), the second most common age related ND. It has been postulated that oligomeric species of aSyn, rather than more mature aggregated forms of the protein, are the causative agents of cytotoxicity. In recent years, we have been investigating the molecular mechanisms underlying the initial steps of aSyn accumulation in living cells. Using an unbiased genome-wide lentiviral RNAi screen we identified trafficking and kinase genes as modulators of aSyn oligomerization, aggregation, and toxicity. Among those, Rab8b, Rab11a, Rab13 and Slp5 were found to promote the clearance of aSyn inclusions and reduce aSyn toxicity. Moreover, we found that endocytic recycling and secretion of aSyn was enhanced upon expression of Rab11a or Rab13 in cells accumulating aSyn inclusions. Altogether, our findings suggest specific trafficking steps may prove beneficial as targets for therapeutic intervention in synucleinopathies, and should be further investigated in other models.

Keywords: Parkinson disease; Rab GTPases; Slp5; aggregation; neuronal trafficking; oligomerization; α-Synuclein.

Figure 1.

Strategy used for the identification of genetic modifiers of aSyn oligomerization in living cells. A human shRNA library targeting trafficking and phosphotransferase genes was screened using a stable cell line expressing aSyn-BiFC constructs, and basal fluorescent levels were determined (central image). Genes modifying aSyn oligomerization by at least 50% (left and right images) were identified using fluorescence microscopy analysis and were considered for further validation.

Figure 2.

Genes found to modulate the accumulation and secretion of different aggregated species of aSyn. To study the effect of the genes tested on different steps of the process of aSyn aggregation, we used cell models of aSyn oligomerization and aggregation (dashed rectangles). In an unbiased large-scale RNAi screen, we identified 4 trafficking genes that modulate aSyn oligomerization, aggregation, toxicity and secretion. Rab8b is involved in polarized vesicular trafficking (endoplasmic reticulum to plasma membrane). Rab11a regulates endocytic recycling pathway and participates specifically in transferrin recycling. Rab13 regulates trans-Golgi network, recycling endosomes and cell/tight junctions. Slp5 is a Rab27a effector protein and plays a role in exosome release. We found that Rab8b, Rab11a and Rab13 overexpression inhibits oligomerization and aggregation of aSyn. Moreover, Rab11a and Rab13 increase aSyn secretion through the recycling endocytic (RE) route only when aSyn inclusions are present in the cell. Slp5 inhibits aSyn aggregation, but not oligomerization, and increases aSyn secretion in the cell model of aSyn oligomerization.