title: Negative regulation of cyclin D3 expression by transcription factor c-Ets1 in umbilical cord hematopoietic cells.
abstract: OBJECTIVE
  To investigate the role of transcription factor c-Ets1 in cyclin D3 expression and its effects on the proliferation of umbilical cord hematopoietic cells.
  
  
  METHODS
  Cyclin D3 promoter deletion constructs were generated and transfected into CD34(+) cells. Dual luciferase reporter assays and TFSEARCH software were used to identify negative regulatory domains and to predict putative transcription factors involved in cyclin D3 downregulation. Expression of c-Ets1 in CD34(+) cells was detected using electrophoretic mobility shift and super shift assays. Point mutants of c-Ets1 binding sites were constructed. The wild-type c-Ets1 and the mutant promoter constructs were co-transfected into CD34(+) cells to determine the promoter activity. The impact of c-Ets1 expression on the proliferation of CD34(+) cells was assessed using MTT assay.
  
  
  RESULTS
  Nine cyclin D3 promoter deletion constructs were generated. A negative regulatory domain containing c-Ets1 binding sites was identified between -439 bp and -362 bp. Transfection of the promoter deletion constructs containing mutant c-Ets1 binding sites enhanced cyclin D3 promoter activity. However, the opposite results were observed when CD34(+) cells were co-transfected with wildtype c-Ets1 and its promoter deletion constructs. The overexpression of c-Ets1 could suppress cyclin D3 mRNA and protein levels. In addition, it inhibits the proliferation of CD34(+) cells.
  
  
  CONCLUSIONS
  c-Ets1 functions as a negative transcription factor, down-regulating the expression of cyclin D3, which leads to inhibition of CD34(+) cell proliferation.
authors.affiliation: Department of Hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
authors.forename: Fan-kai
authors.initials: FK
authors.lastname: Meng
authors.forename: Han-ying
authors.initials: HY
authors.lastname: Sun
authors.forename: Xi-you
authors.initials: XY
authors.lastname: Tan
authors.forename: Chun-rui
authors.initials: CR
authors.lastname: Li
authors.forename: Jian-feng
authors.initials: JF
authors.lastname: Zhou
authors.forename: Wen-li
authors.initials: WL
authors.lastname: Liu
mesh_terms: D001665:Binding Sites; D017252:CD3 Complex / Q000032:analysis; D049109:Cell Proliferation; D002478:Cells, Cultured; D056743:Cyclin D3 / Q000235:genetics*; D005312:Fetal Blood / Q000166:cytology*; D005786:Gene Expression Regulation; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D017354:Point Mutation; D050784:Proto-Oncogene Protein c-ets-1 / Q000235:genetics / Q000378:metabolism*
chemical_list: D017252:CD3 Complex; D056743:Cyclin D3; D050784:Proto-Oncogene Protein c-ets-1