title: MMTV-associated transcription factor binding sites increase nm23-H1 metastasis suppressor gene expression in human breast carcinoma cell lines.
abstract: We hypothesize that elevation of nm23-HI metastasis suppressor gene expression in micrometastatic tumor cells may reduce their subsequent colonization and invasion, and induce differentiation, with a clinical benefit. This report presents the first analysis of the nm23-HI promoter to identify sites which can increase its transcription. Deletion mapping of a 2.1 kb nm23-H1 promoter fragment tethered to a reporter gene identified three regions involved in its differential expression levels among a panel of human breast carcinoma cell lines: a 195 bp NheI-XbaI fragment responsible for basal expression levels, a 248 bp AvrII-Nhel fragment which contributed to the elevated nm23-H1 expression observed in the high expressing cell lines, and a 544 bp AvrII fragment containing an inhibitory element. Examination of the 248 bp AvrII-NheI fragment revealed the unexpected presence of three transcription factor binding sites (MAF/Ets, CTF/NF1 half site and ACAAAG enhancer) previously identified in the MMTV-LTR, and in WAP and milk gene promoters, proposed to mediate mammary-specific gene expression. Mutation of the three sites, individually or together, resulted in two-fold reductions in reporter gene expression. As controls, the same panel of mutations caused a different pattern of reporter gene expression in a non-mammary cell line, and mutation of another nearby site was without effect on nm23-HI. Our data identify a complex regulatory pattern for nm23-H1 transcription, and suggest that a mammary-specific cassette of transcription factors contribute to its elevated expression
authors.affiliation: Women's Cancers Section, Laboratory of Pathology, National Cancer Institute, Bethesda, Maryland 20892, USA. taoufik@mail.nih.gov
authors.forename: Taoufik
authors.initials: T
authors.lastname: Ouatas
authors.forename: Susan E
authors.initials: SE
authors.lastname: Clare
authors.forename: Melanie T
authors.initials: MT
authors.lastname: Hartsough
authors.forename: Abel
authors.initials: A
authors.lastname: De La Rosa
authors.forename: Patricia S
authors.initials: PS
authors.lastname: Steeg
mesh_terms: D001665:Binding Sites; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D022762:CCAAT-Enhancer-Binding Proteins / Q000378:metabolism; D002277:Carcinoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D016384:Consensus Sequence; D004273:DNA, Neoplasm / Q000235:genetics*; D024202:Electrophoretic Mobility Shift Assay; D004742:Enhancer Elements, Genetic / Q000235:genetics; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D017930:Genes, Reporter; D016147:Genes, Tumor Suppressor; D006801:Humans; D008324:Mammary Tumor Virus, Mouse / Q000235:genetics*; D020559:Monomeric GTP-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics; D016296:Mutagenesis; D051839:NFI Transcription Factors; D054778:NM23 Nucleoside Diphosphate Kinases; D009362:Neoplasm Metastasis / Q000235:genetics*; D009363:Neoplasm Proteins / Q000096:biosynthesis* / Q000235:genetics; D009701:Nucleoside-Diphosphate Kinase; D009928:Organ Specificity; D012150:Polymorphism, Restriction Fragment Length; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D050783:Proto-Oncogene Proteins c-ets; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D017384:Sequence Deletion; D020079:Terminal Repeat Sequences / Q000235:genetics*; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
chemical_list: D022762:CCAAT-Enhancer-Binding Proteins; C077305:CTF-1 transcription factor; D004273:DNA, Neoplasm; D051839:NFI Transcription Factors; D054778:NM23 Nucleoside Diphosphate Kinases; D009363:Neoplasm Proteins; D011518:Proto-Oncogene Proteins; D050783:Proto-Oncogene Proteins c-ets; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C517938:NME1 protein, human; C517939:Nme1 protein, mouse; D009701:Nucleoside-Diphosphate Kinase; D020559:Monomeric GTP-Binding Proteins