Journal Article

MMTV-associated transcription factor binding sites increase nm23-H1 metastasis suppressor gene expression in human breast carcinoma cell lines

De La Rosa, Abel, Hartsough, Melanie T., Clare, Susan E., Ouatas, Taoufik, Steeg, Patricia S.

2002

Overview

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

• Publisher: Springer Nature B.V
• Subject: Binding Sites / Breast Neoplasms - genetics / Breast Neoplasms - metabolism / Breast Neoplasms - pathology / Carcinoma - genetics / Carcinoma - metabolism / Carcinoma - pathology / CCAAT-Enhancer-Binding Proteins - metabolism / Consensus Sequence / DNA, Neoplasm - genetics / Electrophoretic Mobility Shift Assay / Enhancer Elements, Genetic - genetics / Female / Gene Expression Regulation, Neoplastic / Genes, Reporter / Genes, Tumor Suppressor / Humans / Mammary Tumor Virus, Mouse - genetics / Monomeric GTP-Binding Proteins - biosynthesis / Monomeric GTP-Binding Proteins - genetics / Mutagenesis / Neoplasm Metastasis - genetics / Neoplasm Proteins - biosynthesis / Neoplasm Proteins - genetics / NFI Transcription Factors / NM23 Nucleoside Diphosphate Kinases / Nucleoside-Diphosphate Kinase / Organ Specificity / Polymorphism, Restriction Fragment Length / Promoter Regions, Genetic - genetics / Proto-Oncogene Proteins - metabolism / Proto-Oncogene Proteins c-ets / Recombinant Fusion Proteins - biosynthesis / Sequence Deletion / Terminal Repeat Sequences - genetics / Transcription Factors - biosynthesis / Transcription Factors - genetics / Transcription Factors - metabolism / Transcription, Genetic / Tumor Cells, Cultured