Tumour metastasis suppressor, nm23-beta, inhibits gelatinase A transcription by interference with transactivator Y-box protein-1 (YB-1).
Gelatinase A transcriptional regulation is the consequence of combinatorial interactions with key promoter and enhancer elements identified within this gene. A potent 40 bp enhancer response element, RE-1, located in the near 5' flanking regions of the rat and human gelatinase A genes drives high-level expression in glomerular mesangial cells (MCs). Southwestern-blot analysis of MC nuclear extracts revealed specific interactions of RE-1 with at least four proteins, of which three have been identified as p53, activator protein 2 and the single-stranded DNA-binding factor Y-box protein-1 (YB-1). In the present study, we report the identification of a fourth 17 kDa RE-1-binding protein as the rat homologue (nm23-beta) of the human nm23-H1 metastasis suppressor gene. Recombinant nm23-beta protein bound only the single-stranded forms of the RE-1 sequence. Mutagenesis revealed direct interaction of nm23-beta with a repeat sequence, 5'-GGGTTT-3', shown previously to specifically interact with YB-1 [Mertens, Harendza, Pollock and Lovett (1997) J. Biol. Chem. 272, 22905-22912], and recombinant nm23-beta protein competed for single-stranded YB-1 binding. Transient transfection of MC with an nm23-beta expression plasmid within the context of a RE-1/simian virus 40 promoter/luciferase reporter yielded a concentration-dependent repression (80-90%) of luciferase activity in MC and Rat1 fibroblasts. A similar pattern of nm23-beta repression was demonstrated within the context of the RE-1/homologous gelatinase A promoter. Co-transfection of nm23-beta blocked YB-1-mediated activation of transcription and expression of gelatinase A. Nm23-beta may be an important physiological regulator of gelatinase A transcription that acts by competitive interference with the single-stranded transactivator YB-1. Gelatinase A is a key mediator of tumour metastasis, suggesting that competitive suppression of transcription by nm23-beta (or the human nm23-H1) may be a component of the reduced metastatic capabilities of cells expressing high levels of this protein.
Cheng
Sunfa
S
Department of Medicine, San Francisco Veterans Affair Medical Center, University of California, San Francisco 94121, USA.
Alfonso-Jaume
Maria Alexandra
MA
Mertens
Peter R
PR
Lovett
David H
DH
D000818:Animals; D001667:Binding, Competitive; D015139:Blotting, Southern; D015153:Blotting, Western; D022762:CCAAT-Enhancer-Binding Proteins / Q000378:metabolism*; D002455:Cell Division; D002467:Cell Nucleus / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D004268:DNA-Binding Proteins; D005347:Fibroblasts / Q000378:metabolism; D017930:Genes, Reporter; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism*; D020559:Monomeric GTP-Binding Proteins / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D051839:NFI Transcription Factors; D054778:NM23 Nucleoside Diphosphate Kinases; D009362:Neoplasm Metastasis; D009687:Nuclear Proteins; D009701:Nucleoside-Diphosphate Kinase / Q000378:metabolism; D009841:Oligonucleotides / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection; D051840:Y-Box-Binding Protein 1
D022762:CCAAT-Enhancer-Binding Proteins; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D051839:NFI Transcription Factors; D054778:NM23 Nucleoside Diphosphate Kinases; D009687:Nuclear Proteins; D009841:Oligonucleotides; D012333:RNA, Messenger; D011994:Recombinant Proteins; D014157:Transcription Factors; D051840:Y-Box-Binding Protein 1; C070856:YBX1 protein, human; C486169:Ybx1 protein, rat; D008156:Luciferases; C517938:NME1 protein, human; D009701:Nucleoside-Diphosphate Kinase; D020778:Matrix Metalloproteinase 2; D020559:Monomeric GTP-Binding Proteins