title: Receptor-interacting protein 140 overexpression promotes neuro-2a neuronal differentiation by ERK1/2 signaling.
abstract: BACKGROUND
  Abnormal neuronal differentiation plays an important role in central nervous system (CNS) development abnormalities such as Down syndrome (DS), a disorder that results directly from overexpression of genes in trisomic cells. Receptor-interacting protein 140 (RIP140) is significantly upregulated in DS brains, suggesting its involvement in DS CNS development abnormalities. However, the role of RIP140 in neuronal differentiation is still not clear. The current study aimed to investigate the effect of RIP140 overexpression on the differentiation of neuro-2a (N2a) neuroblastoma cells, in vitro.
  
  
  METHODS
  Stably RIP140-overexpressing N2a (N2a-RIP140) cells were used as a neurodevelopmental model, and were constructed by lipofection and overexpression validated by real-time polymerase chain reaction and Western blot. Retinoic acid (RA) was used to stimulate N2a differentiation. Combining the expression of Tuj1 at the mRNA and protein levels, the percentage of cells baring neurites, and the number of neurites per cell body was semi-quantified to determine the effect of RIP140 on differentiation of N2a cells. Furthermore, western blot and the ERK1/2 inhibitor U0126 were used to identify the specific signaling pathway by which RIP140 induces differentiation of N2a cells. Statistical significance of the differences between groups was determined by one-way analysis of variance followed by the Dunnett test.
  
  
  RESULTS
  Compared to untransfected N2a cells RIPl40 expression in N2a-RIP140 cells was remarkably upregulated at both the mRNA and protein levels. N2a-RIP140 cells had a signiﬁcantly increased percentage of cells baring neurites, and numbers of neurites per cell, as compared to N2a cells, in the absence and presence of RA (P < 0.05). In addition, Tuj1, a neuronal biomarker, was strongly upregulated in N2a-RIP140 cells (P < 0.05) and phosphorylated ERK1/2 (p-ERK1/2) levels in N2a-RIP140 cells were dramatically increased, while differentiation was inhibited by the ERK1/2-specific inhibitor U0126.
  
  
  CONCLUSIONS
  RIP140 overexpression promotes N2a cell neuronal differentiation by activating the ERK1/2 pathway.
authors.forename: Xiao
authors.initials: X
authors.lastname: Feng
authors.forename: Weidong
authors.initials: W
authors.lastname: Yu
authors.forename: Rong
authors.initials: R
authors.lastname: Liang
authors.forename: Cheng
authors.initials: C
authors.lastname: Shi
authors.forename: Zhuran
authors.initials: Z
authors.lastname: Zhao
authors.affiliation: Department of Pediatrics, Peking University People's Hospital, Beijing 100044, China.
authors.forename: Jingzhu
authors.initials: J
authors.lastname: Guo
mesh_terms: D015153:Blotting, Western; D002454:Cell Differentiation / Q000502:physiology; D002460:Cell Line; D006801:Humans; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D056971:Nuclear Receptor Co-Repressor 1 / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology
chemical_list: D056971:Nuclear Receptor Co-Repressor 1; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3