title: [Transcriptional activation of TMSG-1 by complex of KLF6 and Sp1].
abstract: OBJECTIVE
  To investigate the regulatory mechanism of the transcription of tumor metastasis suppressor gene TMSG-1.
  
  
  METHODS
  Luciferase reporter assay and site-directed mutagenesis were used to analyze the regulatory region of TMSG-1. Electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) were carried out to verify the interaction of KLF6 and Sp1 with the regulatory region of TMSG-1. Co-immunoprecipitation (CoIP) was performed to analyze the interaction between KLF6 and Sp1. TMSG-1 and wt-KLF6 mRNA expressions in cells with different metastatic capacities were quantitated by real-time PCR. Cell invasive capability was determined by Matrigel invasion assay.
  
  
  RESULTS
  A 63 bp inducible regulatory region (+59 bp - +123 bp) in exon 1 was identified by luciferase assay using reporter plasmids with a series of TMSG-1 regulatory region deletions. Mutations in KLF6/Sp1 binding sites of this region resulted in a decrease of luciferase activity, while cotransfection with KLF6 or Sp1 expressing plasmids led to a remarkable increase of luciferase activity. EMSA and ChIP demonstrated that KLF6 as well as Sp1 interacted with this region. CoIP also indicated a possible interaction between KLF6 and Sp1 proteins. In the highly metastatic cell sublines, a low level of wild type KLF6 was associated synchronously with a low TMSG-1 level. Prostate carcinoma cells overexpressing KLF6 exhibited a higher TMSG-1 level and a lower invasive capability.
  
  
  CONCLUSIONS
  Transcription factor complex of KLF6 and Sp1 may participate in the inducible transcriptional regulation of TMSG-1, and a decreased wild type KLF6 expression is likely associated with a low TMSG-1 level in the highly metastatic cell sublines.
authors.affiliation: Department of Pathology, Peking University, Beijing, China.
authors.forename: Miao-zi
authors.initials: MZ
authors.lastname: Gong
authors.forename: Jiang-feng
authors.initials: JF
authors.lastname: You
authors.forename: Fei
authors.initials: F
authors.lastname: Pei
authors.forename: Xiang-lin
authors.initials: XL
authors.lastname: Cui
authors.forename: Gang
authors.initials: G
authors.lastname: Li
authors.forename: Jie
authors.initials: J
authors.lastname: Zheng
mesh_terms: D001665:Binding Sites / Q000235:genetics; D045744:Cell Line, Tumor; D024202:Electrophoretic Mobility Shift Assay; D006801:Humans; D047468:Immunoprecipitation; D000075723:Kruppel-Like Factor 6; D051741:Kruppel-Like Transcription Factors / Q000235:genetics / Q000378:metabolism*; D008175:Lung Neoplasms / Q000378:metabolism / Q000473:pathology; D008297:Male; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009361:Neoplasm Invasiveness; D011471:Prostatic Neoplasms / Q000378:metabolism / Q000473:pathology; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D051049:Sphingosine N-Acyltransferase / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation; D014162:Transfection; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: C105465:KLF6 protein, human; D000075723:Kruppel-Like Factor 6; D051741:Kruppel-Like Transcription Factors; D008565:Membrane Proteins; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D016329:Sp1 Transcription Factor; D025521:Tumor Suppressor Proteins; C438311:CERS2 protein, human; D051049:Sphingosine N-Acyltransferase