title: Id2 promotes the invasive growth of MCF-7 and SKOV-3 cells by a novel mechanism independent of dimerization to basic helix-loop-helix factors.
abstract: BACKGROUND
  Inhibitor of differentiation 2 (Id2) is a critical factor for cell proliferation and differentiation in normal vertebrate development. Most of the biological function of Id2 has been ascribed to its helix-loop-helix motif. Overexpression of Id2 is frequently observed in various human tumors, but its role for invasion potential in tumor cells is dispute. We aimed to reveal the role of Id2 in invasion potential in poorly invasive and estrogen receptor alpha (ERalpha)-positive MCF-7 and SKOV-3 cancer cells.
  
  
  METHODS
  MCF-7 and SKOV-3 cells were stably transfected with the wild-type, degradation-resistant full-length or helix-loop-helix (HLH)-deleted Id2, respectively. Protein levels of Id2 and its mutants and E-cadherin were determined by western blot analysis and mRNA levels of Id2 and its mutants were determined by RT-PCR. The effects of Id2 and its mutants on cell proliferation were determined by [3H]-thymidine incorporation assay and the 3- [4, 5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) dye method. The in vitro invasion potential of cells was evaluated by Transwell assay. Cell motility was assessed by scratch wound assay. The promoter activity of E-cadherin was determined by cotransfection and luciferase assays.
  
  
  RESULTS
  Ectopic transfection of the wild-type Id2 markedly increased the protein and mRNA expression of Id2 in MCF-7 and SKOV-3 cells; the protein level but not mRNA level was further increased by transfection with the degradation-resistant Id2 form. The ectopic expression of Id2 or its mutants did not alter proliferation of either MCF-7 or SKOV-3 cells. Transfection of the wild-type Id2 significantly induced the invasion potential and migratory capacity of cells, which was further augmented by transfection with the degradation-resistant full-length or HLH-deleted Id2. E-cadherin protein expression and transactivation of the proximal E-cadherin promoter were markedly suppressed by the degradation-resistant full-length or HLH-deleted Id2 but not wild-type Id2. Ectopic expression of E-cadherin in MCF-7 and SKOV-3 cells only partially blunted the invasion potential induced by the degradation-resistant HLH-deleted Id2.
  
  
  CONCLUSIONS
  Overexpression of Id2 in ERalpha-positive epithelial tumor cells indeed increases the cells' invasive potential through a novel mechanism independent of dimerization to basic helix-loop-helix factors. E-cadherin contributes only in part to Id2-induced cell invasion when Id2 is accumulated to a higher level in some specific cell types.
authors.affiliation: Department of Molecular Biology, Institute of Basic Medicine, Chinese PLA General Hospital, 28 Fu Xing Road, Beijing, 100853, PR China. mengyg6512@vipsina.com
authors.forename: Yuanguang
authors.initials: Y
authors.lastname: Meng
authors.forename: Chenglei
authors.initials: C
authors.lastname: Gu
authors.forename: Zhiqiang
authors.initials: Z
authors.lastname: Wu
authors.forename: Yali
authors.initials: Y
authors.lastname: Zhao
authors.forename: Yiling
authors.initials: Y
authors.lastname: Si
authors.forename: Xiaobing
authors.initials: X
authors.lastname: Fu
authors.forename: Weidong
authors.initials: W
authors.lastname: Han
mesh_terms: D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D015820:Cadherins / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics; D048708:Cell Growth Processes / Q000502:physiology; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000502:physiology; D019281:Dimerization; D047628:Estrogen Receptor alpha / Q000096:biosynthesis; D005260:Female; D018257:Helix-Loop-Helix Motifs; D006801:Humans; D051797:Inhibitor of Differentiation Protein 2 / Q000096:biosynthesis* / Q000235:genetics; D009361:Neoplasm Invasiveness; D010051:Ovarian Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D014158:Transcription, Genetic; D014162:Transfection
chemical_list: D015820:Cadherins; D047628:Estrogen Receptor alpha; C497450:ID2 protein, human; D051797:Inhibitor of Differentiation Protein 2; D012333:RNA, Messenger; D034741:RNA, Small Interfering