Nurr1 is phosphorylated by ERK2 in vitro and its phosphorylation upregulates tyrosine hydroxylase expression in SH-SY5Y cells.
Nurr1 is an orphan nuclear receptor essential for development and survival of dopaminergic neurons. Mutations in Nurr1 are associated with Parkinson's disease (PD) and there is a correlation between Nurr1 and tyrosine hydroxylase (TH) expression in PD brain. Two domains, activation function 1 (AF1) at the N-terminus and AF2 at the C-terminus of Nurr1, are important for Nurr1 activation. AF1 domain is conserved in NGFI-B/Nurr1/Nor-1 family members and MAPK signal pathway is involved in AF1 activity. Using in vitro phoshorylation assays, we have shown that ERK2 is a kinase to phosphorylate Nurr1 on multiple sites. S126 and T132, which are located near AF1 core of Nurr1, are dominant sites phosphorylated by ERK2. Moreover, using GST pull-down and co-IP assays, we identified that both the N-terminus of Nurr1 containing three ERK docking domains and another ERK docking domain in Nurr1 DNA binding domain are able to bind to ERK2. Furthermore, overexpression of a constitutively active form of MEK1, together with Nurr1 and mouse ERK2, greatly increases the tyrosine hydroxylase expression in SH-SY5Y cells. Reporter gene assays show that Nurr1Delta124-133/T185A, an ERK2 phospho-site mutant form, could not further increase its transcriptional activity on TH promoter, suggesting that Nurr1 phosphorylation by ERK2 may regulate its transcriptional activity on TH promoter. Thus, our results indicate that Nurr1 phosphorylation by ERK2 may play a role in regulating the TH expression.
Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science & Technology of China, Hefei, Anhui 230027, China.
Tao
T
Zhang
Nali
N
Jia
Erkang
E
Fei
Pingping
P
Wang
Zhandi
Z
Liao
Lili
L
Ding
Ming
M
Yan
Nobuyuki
N
Nukina
Jiangning
J
Zhou
Guanghui
G
Wang
D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D005786:Gene Expression Regulation; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D066298:In Vitro Techniques; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D009369:Neoplasms; D057126:Nuclear Receptor Subfamily 4, Group A, Member 2; D010766:Phosphorylation; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection; D014446:Tyrosine 3-Monooxygenase / Q000096:biosynthesis*; D015854:Up-Regulation
D004268:DNA-Binding Proteins; C539882:NR4A2 protein, human; C539883:Nr4a2 protein, mouse; D057126:Nuclear Receptor Subfamily 4, Group A, Member 2; D014157:Transcription Factors; D014446:Tyrosine 3-Monooxygenase; D019950:Mitogen-Activated Protein Kinase 1