title: [The effect of PKC phosphorylation sites mutation in JWA coding region on TPA-induced MCF-7 cell differentiation].
abstract: OBJECTIVE
  To investigate the effect of PKC phosphorylation sites mutation in JWA coding region on TPA-induced MCF-7 cell differentiation.
  
  
  METHODS
  Site directed gene mutation was used to construct one or two PKC sites mutations in pEGFP-N1-JWA vectors, and transfected into MCF-7 cells by polyfect reagent, and cell differentiation was characterized by accumulation of lipid droplet as indicated by positive Oil-red-O staining of cells.
  
  
  RESULTS
  All these transfected cell lines, MCF-7-N1(transfected with pEGFP-N1 vector), MCF-7-JWA(transfected with pEGFP-N1-JWA vector), MCF-7-JWA-1(transfected with PKC site 1 mutation pEGFP-N1-JWA vector), MCF-7-JWA-2(transfected with PKC site 2 mutation pEGFP-N1-JWA vector), MCF-7-JWA-1+2 (transfected with both PKC site 1 and 2 mutation pEGFP-N1-JWA vector) were treated with 20 nmol/L TPA for 48 h, and the percentages of positive Oil-red-O staining of cells were 48%, 67%, 69%, 67% and 70% respectively. The percentages of cell differentiation in JWA containing vectors transfected cells treated with TPA were significantly higher those of MCF-7-N1 cells (vector only control). However there were no significant differences between mutated and unmutated cells.
  
  
  CONCLUSIONS
  JWA transfection enhanced MCF-7 cell differentiation induced by TPA significantly, and PKC sites mutation in JWA coding region has no obviously effect on TPA-induced MCF-7 cell differentiation.
authors.affiliation: School of Public Health, Nanjing Medical University, Nanjing 210029, China.
authors.forename: Jian
authors.initials: J
authors.lastname: Ye
authors.forename: Ai-ping
authors.initials: AP
authors.lastname: Li
authors.forename: Wen
authors.initials: W
authors.lastname: Qiu
authors.forename: Jin-hong
authors.initials: JH
authors.lastname: Zhou
authors.forename: Jian-wei
authors.initials: JW
authors.lastname: Zhou
mesh_terms: D002454:Cell Differentiation / Q000187:drug effects*; D045744:Cell Line, Tumor; D005260:Female; D005822:Genetic Vectors; D006801:Humans; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D017354:Point Mutation; D011493:Protein Kinase C / Q000235:genetics* / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014162:Transfection
chemical_list: D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate