Mammalian ribosomal and chaperone protein RPS3A counteracts α-synuclein aggregation and toxicity in a yeast model system

摘要

Accumulation of aggregated forms of αSyn (α-synuclein) into Lewy bodies is a known hallmark associated with neuronal cell death in Parkinson's disease. When expressed in the yeast Saccharomyces cerevisiae , αSyn interacts with the plasma membrane, forms inclusions and causes a concentration-dependent growth defect. We have used a yeast mutant, cog6 Δ, which is particularly sensitive to moderate αSyn expression, for screening a mouse brain-specific cDNA library in order to identify mammalian proteins that counteract αSyn toxicity. The mouse ribosomal and chaperone protein RPS3A was identified as a suppressor of αSyn [WT (wild-type) and A53T] toxicity in yeast. We demonstrated that the 50 N-terminal amino acids are essential for this function. The yeast homologues of RPS3A were not effective in suppressing the αSyn-induced growth defect, illustrating the potential of our screening system to identify modifiers that would be missed using yeast gene overexpression as the first screening step. Co-expression of mouse RPS3A delayed the formation of αSyn–GFP inclusions in the yeast cells. The results of the present study suggest that the recently identified extraribosomal chaperonin function of RPS3A also acts on the neurodegeneration-related protein αSyn and reveal a new avenue for identifying promising candidate mammalian proteins involved in αSyn functioning.Abbreviations: COG6, component of oligomeric Golgi complex 6; ER, endoplasmic reticulum; 5-FOA, 5-fluoro-orotic acid; Gapdh, glyceraldehyde-3-phosphate dehydrogenase; HBx, hepatitis B protein X; HRP, horseradish peroxidase; LB, Lewy body; MPTP, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine; NA, numerical aperture; PD, Parkinson’s disease; Pgk1, phosphoglycerate kinase 1; PSMA2, proteasome subunit α type?2; Spir1CT, C-terminus of Spire homologue 1; RPS3A, ribosomal protein S3A; SC, synthetic complete; SD, synthetic dextrose; SNARE, soluble N-ethylmaleimide-sensitive fusion protein-attachment protein receptor; αSyn, α-synuclein; WT, wild-type; yeGFP, yeast-enhanced GFP; YPD, yeast extract/peptone/dextrose