Limits...
Nigral overexpression of alpha-synuclein in the absence of parkin enhances alpha-synuclein phosphorylation but does not modulate dopaminergic neurodegeneration.

Van Rompuy AS, Oliveras-Salvá M, Van der Perren A, Corti O, Van den Haute C, Baekelandt V - Mol Neurodegener (2015)

Bottom Line: Conflicting results were reported about the effect of the absence of parkin on alpha-synuclein-mediated neurotoxicity using a transgenic approach.The increased alpha-synuclein phosphorylation was confirmed in a parkin knockdown cell line.These findings support a functional relationship between parkin and alpha-synuclein phosphorylation in rodent brain.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Neurobiology and Gene Therapy, Department of Neurosciences, KU Leuven, Flanders, Belgium. annexsophie.vanrompuy@uzleuven.be.

ABSTRACT

Background: Alpha-synuclein is a key protein in the pathogenesis of Parkinson's disease. Mutations in the parkin gene are the most common cause of early-onset autosomal recessive Parkinson's disease, probably through a loss-of-function mechanism. However, the molecular mechanism by which loss of parkin function leads to the development of the disease and the role of alpha-synuclein in parkin-associated Parkinson's disease is still not elucidated. Conflicting results were reported about the effect of the absence of parkin on alpha-synuclein-mediated neurotoxicity using a transgenic approach. In this study, we investigated the effect of loss of parkin on alpha-synuclein neuropathology and toxicity in adult rodent brain using viral vectors. Therefore, we overexpressed human wild type alpha-synuclein in the substantia nigra of parkin knockout and wild type mice using two different doses of recombinant adeno-associated viral vectors.

Results: No difference was observed in nigral dopaminergic cell loss between the parkin knockout mice and wild type mice up to 16 weeks after viral vector injection. However, the level of alpha-synuclein phosphorylated at serine residue 129 in the substantia nigra was significantly increased in the parkin knockout mice compared to the wild type mice while the total expression level of alpha-synuclein was similar in both groups. The increased alpha-synuclein phosphorylation was confirmed in a parkin knockdown cell line.

Conclusions: These findings support a functional relationship between parkin and alpha-synuclein phosphorylation in rodent brain.

No MeSH data available.


Related in: MedlinePlus

Increased P-S129 α-SYN in human SHSY5Y neuroblastoma cells after parkin knockdown. Western blotting against PLK2, parkin, PP2A, P-Ser129 α-SYN and α-SYN. The miRs against parkin induced both parkin knockdown and an increase in P-Ser129 α-SYN signal without affecting α-SYN, PLK2 and PP2A levels. A miR against firefly luciferase (Fluc) was used as control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4477319&req=5

Fig7: Increased P-S129 α-SYN in human SHSY5Y neuroblastoma cells after parkin knockdown. Western blotting against PLK2, parkin, PP2A, P-Ser129 α-SYN and α-SYN. The miRs against parkin induced both parkin knockdown and an increase in P-Ser129 α-SYN signal without affecting α-SYN, PLK2 and PP2A levels. A miR against firefly luciferase (Fluc) was used as control.

Mentions: To investigate the mechanism behind this increased α-SYN phosphorylation in the absence of parkin we induced stable parkin knockdown using microRNA (miR)-based lentiviral vectors in human SHSY5Y neuroblastoma cells overexpressing WT human α-SYN. Two miRparkin lentiviral vectors induced efficient knockdown of parkin as shown by Q-RT-PCR (data not shown) and Western blotting (Figure 7). In agreement with the in vivo data we observed an increase of α-SYN phosphorylation at serine residue 129 without affecting the total α-SYN levels in cell culture (Figure 7). Since Polo-Like-Kinase-2 (PLK2) was shown to phosphorylate α-SYN at S129 [36] and protein phosphatase-2A (PP2A) is a known α-SYN phosphatase at S129 [37], we verified the levels of PLK2 and PP2A in both parkin knockdown and the control cell lines but no differences were observed  in expression of either protein (Figure 7).Figure 7


Nigral overexpression of alpha-synuclein in the absence of parkin enhances alpha-synuclein phosphorylation but does not modulate dopaminergic neurodegeneration.

Van Rompuy AS, Oliveras-Salvá M, Van der Perren A, Corti O, Van den Haute C, Baekelandt V - Mol Neurodegener (2015)

Increased P-S129 α-SYN in human SHSY5Y neuroblastoma cells after parkin knockdown. Western blotting against PLK2, parkin, PP2A, P-Ser129 α-SYN and α-SYN. The miRs against parkin induced both parkin knockdown and an increase in P-Ser129 α-SYN signal without affecting α-SYN, PLK2 and PP2A levels. A miR against firefly luciferase (Fluc) was used as control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4477319&req=5

Fig7: Increased P-S129 α-SYN in human SHSY5Y neuroblastoma cells after parkin knockdown. Western blotting against PLK2, parkin, PP2A, P-Ser129 α-SYN and α-SYN. The miRs against parkin induced both parkin knockdown and an increase in P-Ser129 α-SYN signal without affecting α-SYN, PLK2 and PP2A levels. A miR against firefly luciferase (Fluc) was used as control.
Mentions: To investigate the mechanism behind this increased α-SYN phosphorylation in the absence of parkin we induced stable parkin knockdown using microRNA (miR)-based lentiviral vectors in human SHSY5Y neuroblastoma cells overexpressing WT human α-SYN. Two miRparkin lentiviral vectors induced efficient knockdown of parkin as shown by Q-RT-PCR (data not shown) and Western blotting (Figure 7). In agreement with the in vivo data we observed an increase of α-SYN phosphorylation at serine residue 129 without affecting the total α-SYN levels in cell culture (Figure 7). Since Polo-Like-Kinase-2 (PLK2) was shown to phosphorylate α-SYN at S129 [36] and protein phosphatase-2A (PP2A) is a known α-SYN phosphatase at S129 [37], we verified the levels of PLK2 and PP2A in both parkin knockdown and the control cell lines but no differences were observed  in expression of either protein (Figure 7).Figure 7

Bottom Line: Conflicting results were reported about the effect of the absence of parkin on alpha-synuclein-mediated neurotoxicity using a transgenic approach.The increased alpha-synuclein phosphorylation was confirmed in a parkin knockdown cell line.These findings support a functional relationship between parkin and alpha-synuclein phosphorylation in rodent brain.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Neurobiology and Gene Therapy, Department of Neurosciences, KU Leuven, Flanders, Belgium. annexsophie.vanrompuy@uzleuven.be.

ABSTRACT

Background: Alpha-synuclein is a key protein in the pathogenesis of Parkinson's disease. Mutations in the parkin gene are the most common cause of early-onset autosomal recessive Parkinson's disease, probably through a loss-of-function mechanism. However, the molecular mechanism by which loss of parkin function leads to the development of the disease and the role of alpha-synuclein in parkin-associated Parkinson's disease is still not elucidated. Conflicting results were reported about the effect of the absence of parkin on alpha-synuclein-mediated neurotoxicity using a transgenic approach. In this study, we investigated the effect of loss of parkin on alpha-synuclein neuropathology and toxicity in adult rodent brain using viral vectors. Therefore, we overexpressed human wild type alpha-synuclein in the substantia nigra of parkin knockout and wild type mice using two different doses of recombinant adeno-associated viral vectors.

Results: No difference was observed in nigral dopaminergic cell loss between the parkin knockout mice and wild type mice up to 16 weeks after viral vector injection. However, the level of alpha-synuclein phosphorylated at serine residue 129 in the substantia nigra was significantly increased in the parkin knockout mice compared to the wild type mice while the total expression level of alpha-synuclein was similar in both groups. The increased alpha-synuclein phosphorylation was confirmed in a parkin knockdown cell line.

Conclusions: These findings support a functional relationship between parkin and alpha-synuclein phosphorylation in rodent brain.

No MeSH data available.


Related in: MedlinePlus