alpha-synuclein is required for the fibrillar nature of ubiquitinated inclusions induced by proteasomal inhibition in primary neurons.
Proteasomal dysfunction may underlie certain neuro-degenerative conditions such as Parkinson disease. We have shown that pharmacological inhibition of the proteasome in cultured neuronal cells leads to apoptotic death and formation of cytoplasmic ubiquitinated inclusions. These inclusions stain for alpha-synuclein and assume a fibrillar structure, as assessed by thioflavine S staining, and therefore resemble Lewy bodies. alpha-Synuclein is thought to be a central component of Lewy bodies. Whether alpha-synuclein is required for inclusion formation or apoptotic death has not been formally assessed. The present study examines whether alpha-synuclein deficiency in neurons alters their sensitivity to proteasomal inhibition-induced apoptosis or inclusion formation. Cortical neurons derived from alpha-synuclein-null mice showed a similar sensitivity to death induced by the proteasomal inhibitor lactacystin compared with neurons derived from wild-type mice. Furthermore, the absence of alpha-synuclein did not influence the percentage of lactacystin-treated neurons harboring cytoplasmic ubiquitinated inclusions or alter the solubility of such inclusions. In contrast, however, ubiquitinated inclusions in alpha-synuclein-deficient neurons lacked amyloid-like fibrillization, as determined by thioflavine S staining. This indicates that although alpha-synuclein deficiency does not affect the formation of ubiquitinated inclusions, it does significantly alter their structure. The lack of effect on survival in alpha-synuclein knock-out cultures further suggests that the fibrillar nature of the inclusions does not contribute to neuronal degeneration in this model.
Departments of Neurology and Pathology, Columbia University, New York, New York 10032, USA. HR227@columbia.edu
Hardy J
HJ
Rideout
Paula
P
Dietrich
Qiaohong
Q
Wang
William T
WT
Dauer
Leonidas
L
Stefanis
D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis; D052160:Benzothiazoles; D015153:Blotting, Western; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism; D005838:Genotype; D016631:Lewy Bodies / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D008856:Microscopy, Fluorescence; D009419:Nerve Tissue Proteins / Q000378:metabolism / Q000502:physiology*; D009474:Neurons / Q000378:metabolism*; D011480:Protease Inhibitors / Q000494:pharmacology; D061988:Proteasome Inhibitors; D051381:Rats; D051843:Synucleins; D013844:Thiazoles / Q000378:metabolism; D013997:Time Factors; D025801:Ubiquitin / Q000378:metabolism*; D051844:alpha-Synuclein
D052160:Benzothiazoles; D009419:Nerve Tissue Proteins; D011480:Protease Inhibitors; D061988:Proteasome Inhibitors; C497605:Snca protein, mouse; C497606:Snca protein, rat; D051843:Synucleins; D013844:Thiazoles; D025801:Ubiquitin; D051844:alpha-Synuclein; C067713:lactacystin; C009462:thioflavin T; D000111:Acetylcysteine