Proteasome inhibition and aggregation in Parkinson's disease: a comparative study in untransfected and transfected cells.
Dysfunction of the ubiquitin-proteasome system (UPS) has been implicated in Parkinson's disease (PD) and other neurodegenerative disorders. We have investigated the effect of UPS inhibition on the metabolism of alpha-synuclein (SYN) and parkin, two proteins genetically and histopathologically associated to PD. Pharmacological inhibition of proteasome induced accumulation of both parkin and SYN in transfected PC12 cells. We found that this effect was caused by increased protein synthesis rather than impairment of protein degradation, suggesting that inhibition of the UPS might lead to non-specific up-regulation of cytomegalovirus (CMV)-driven transcription. To investigate whether endogenous parkin and SYN can be substrate of the UPS, untransfected PC12 cells and primary mesencephalic neurones were exposed to proteasome inhibitors, and parkin and SYN expression was evaluated at both protein and mRNA level. Under these conditions, we found that proteasome inhibitors did not affect the level of endogenous parkin and SYN. However, we confirmed that dopaminergic neurones were selectively vulnerable to the toxicity of proteasome inhibitors. Our results indicate that studies involving the use of proteasome inhibitors, particularly those in which proteins are expressed from a heterologous promoter, are subjected to potential artefacts that need to be considered for the interpretation of the role of UPS in PD pathogenesis.
Biasini
Emiliano
E
Department of Neuroscience, Istituto di Ricerche Farmacologiche Mario Negri Milano, Italy.
Fioriti
Luana
L
Ceglia
Ilaria
I
Invernizzi
Roberto
R
Bertoli
Alessandro
A
Chiesa
Roberto
R
Forloni
Gianluigi
G
D000818:Animals; D002449:Cell Aggregation / Q000187:drug effects / Q000502:physiology; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007976:Leupeptins / Q000494:pharmacology; D008636:Mesencephalon / Q000187:drug effects / Q000378:metabolism; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D016716:PC12 Cells; D010300:Parkinson Disease / Q000201:enzymology* / Q000235:genetics; D046988:Proteasome Endopeptidase Complex; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D051843:Synucleins; D014162:Transfection / Q000379:methods*; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism; D051844:alpha-Synuclein
D007976:Leupeptins; D009097:Multienzyme Complexes; D009419:Nerve Tissue Proteins; D012333:RNA, Messenger; C497604:SNCA protein, human; C497606:Snca protein, rat; D051843:Synucleins; D051844:alpha-Synuclein; D044767:Ubiquitin-Protein Ligases; C111567:parkin protein; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde