Yeast reveals similar molecular mechanisms underlying alpha- and beta-synuclein toxicity

Abstract

Synucleins belong to a family of intrinsically unstructured proteins that includes alpha-synuclein (aSyn), beta-synuclein (bSyn) and gamma-synuclein (gSyn). aSyn is the most studied member of the synuclein family due to its central role in genetic and sporadic forms of Parkinson's disease and other neurodegenerative disorders known as synucleionopathies. In contrast, bSyn and gSyn have been less studied, but recent reports also suggest that, unexpectedly, these proteins may also cause neurotoxicity. Here, we explored the yeast toolbox to investigate the cellular effects of bSyn and gSyn. We found that bSyn is toxic and forms cytosolic inclusions that are similar to those formed by aSyn. Moreover, we found that bSyn shares similar toxicity mechanisms with aSyn, including vesicular trafficking impairment and induction of oxidative stress. We demonstrate that co-expression of aSyn and bSyn exacerbates cytotoxicity, due to increased dosage of toxic synuclein forms, and that they are able to form heterodimers in both yeast and in human cells. In contrast, gSyn is not toxic and does not form inclusions in yeast cells. Altogether, our findings shed light into the question of whether bSyn can exert toxic effects and confirms the occurrence of aSyn/bSyn heterodimers, opening novel perspectives for the development of novel strategies for therapeutic intervention in synucleinopathies.

A pH-dependent switch promotes β-synuclein fibril formation via glutamate residues

“…They indicate that although ␤S may serve in a neuroprotective role at cytoplasmic pH by preventing ␣S aggregation, this inhibition may be relieved by a switch into its own fibrillation in more acidic organelles, like the lysosome, which enhance protein clearance and enable homeostasis. Several recent reports suggest that ␤S may contribute to a toxic gainof-function in different cellular organelles in a different way than ␣S (16,17). For instance, defects associated with ␤S are linked with defects in endoplasmic reticulum-Golgi-associated proteins in yeast, which is a mildly acidic subcellular environment.…”

“…Furthermore, despite the high sequence similarity, ␤S itself does not form fibrils in vitro at cytoplasmic pH without facilitating agents (15). However, recent reports highlight a role for a possible toxic gain-of-function of ␤S in two different model systems (16,17), and ␤S is a component of vesicle-like lesions in the hippocampus, whose formation accompanies dementia in PD (18). In addition, two ␤S mutations, V70M and P123H, were found in sporadic and familial dementia with LBs and have been suggested to be involved in lysosomal pathology (19 -21).…”

The effects of the novel A53E alpha-synuclein mutation on its oligomerization and aggregation

“…For all experiments, an inoculum was prepared to obtain cells in log growth phase, using synthetic complete (SC) medium [0.67% (w/v) yeast nitrogen base without amino acids (Difco), 1% (w/v) raffinose and 0.79 g.L −1 complete supplement mixture (CSM) (QBiogene)], 200 rpm, 30 °C, as we described [57]. To induce aSyn expression, in liquid medium, cells were grown (OD 600 nm 0.2) in SC selective medium 1% (w/v) galactose (aSyn ON) for 7 h at 30 °C, 200 rpm.…”

“…aSyn cytotoxicity was evaluated by spotting assays. OD 600 nm was set to 0.1 ± 0.005 and 1:10 serially dilutions of each sample were prepared [57]. Then, 4 μL of each dilution was spotted in solid SC selective medium containing 2% glucose (aSyn OFF) or 1% (w/v) galactose (aSyn ON) and incubated at 30 °C for 36–42 h. Images were acquired using ChemiDoc Touch (Bio-Rad).…”

Estudo dos efeitos da superexpressão da alfa-sinucleína sobre o tráfego mitocondrial e autofagia em leveduras, células SH-SY5Y e neurônios dopaminérgicos derivados de hiPSC de pacientes com doença de Parkinson

“…Yeast (Saccharomyces cerevisiae) has been successfully used to investigate alpha-synuclein toxicity related to ER stress, mitochondria, and autophagy dysfunction (Tenreiro et al, 2016;Delorme-Axford et al, 2015;. In other to investigate the role of Miro1, Rab5, and LC3 in alpha-synuclein toxicity, we create a model Δ (knockout) for each of these genes in yeast also expressing A30P or A53T alpha-synuclein.…”