Parkin represses 6-hydroxydopamine-induced apoptosis via stabilizing scaffold protein p62 in PC12 cells.

Abstract

AIM:

Parkin has been shown to exert protective effects against 6-hydroxydopamine (6-OHDA)-induced neurotoxicity in different models of Parkinson disease. In the present study we investigated the molecular mechanisms underlying the neuroprotective action of parkin in vitro.

METHODS:

HEK293, HeLa and PC12 cells were transfected with parkin, parkin mutants, p62 or si-p62. Protein expression and ubiquitination were assessed using immunoblot analysis. Immunoprecipitation assay was performed to identify the interaction between parkin and scaffold protein p62. PC12 and SH-SY5Y cells were treated with 6-OHDA (200 μmol/L), and cell apoptosis was detected using PI and Hoechst staining.

RESULTS:

In HEK293 cells co-transfected with parkin and p62, parkin was co-immunoprecipitated with p62, and parkin overexpression increased p62 protein levels. In parkin-deficient HeLa cells, transfection with wild-type pakin, but not with ligase activity-deficient pakin mutants, significantly increased p62 levels, suggesting that parkin stabilized p62 through its E3 ligase activity. Transfection with parkin or p62 significantly repressed ERK1/2 phosphorylation in HeLa cells, but transfection with parkin did not repress ERK1/2 phosphorylation in p62-knockdown HeLa cells, suggesting that p62 was involved in parkin-induced inhibition on ERK1/2 phosphorylation. Overexpression of parkin or p62 significantly repressed 6-OHDA-induced ERK1/2 phosphorylation in PC12 cells, and parkin overexpression inhibited 6-OHDA-induced apoptosis in PC12 and SH-SY5Y cells.

CONCLUSION:

Parkin protects PC12 cells against 6-OHDA-induced apoptosis via ubiquitinating and stabilizing scaffold protein p62, and repressing ERK1/2 activation.

• Authors: Xiao-ou Hou, Jian-min Si, Hai-gang Ren, Dong Chen, Hong-feng Wang, Zheng Ying, Qing-song Hu, Feng Gao, Guang-hui Wang
• Journal: Acta pharmacologica Sinica (Acta Pharmacol Sin) Vol. 36 Issue 11 Pg. 1300-7 (Nov 2015) ISSN: 1745-7254 [Electronic] United States
• PMID: 26364802 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Heat-Shock Proteins
• Sequestosome-1 Protein
• Sqstm1 protein, rat
• Oxidopamine
• Ubiquitin-Protein Ligases
• parkin protein
• Mitogen-Activated Protein Kinase 1
• Mitogen-Activated Protein Kinase 3

Topics

• Animals
• Apoptosis
• HEK293 Cells
• HeLa Cells
• Heat-Shock Proteins (metabolism)
• Humans
• Mitogen-Activated Protein Kinase 1 (metabolism)
• Mitogen-Activated Protein Kinase 3 (metabolism)
• Oxidopamine (metabolism)
• PC12 Cells
• Rats
• Sequestosome-1 Protein
• Ubiquitin-Protein Ligases (metabolism)
• Ubiquitination