podophyllin and 1-10-phenanthroline

The metal-dependent activation of metallothionein (MT) genes requires the interaction of positive trans-activators (MRFs) with metal-regulatory (MRE) regions of MT promoters. In this report, we examined the role of transition metals in modulating the MRE-binding activities of two different MRE-binding proteins: the metal-regulated factor ZiRF1 and the basal factor SP1. We showed the ability of both proteins to interact with a similar sequence specificity with the cognate target site (MRE-S) of another known MRE-binding protein, mMTF1. We next evaluated the role of metal ions in modulating the MRE-binding activity of recombinant ZiRF1 and basal SP1 proteins by measuring the effect of different metal chelators on DNA interaction. We observed a dose-dependent inhibition of the GST-ZiRF1/MRE-binding activity using three different metal chelators: EDTA, 1,10 PHE and TPEN. Interestingly, EDTA treatment failed to inhibit the recombinant SP1 MRE-binding activity while the effect of 1,10 PHE was comparable to that obtained analyzing 1,10 PHE-treated GST-ZiRF1. The MRE-binding complexes detected in cell extracts showed a response to metal chelator treatment very similar to that displayed by the recombinant ZiRF1 and SP1 proteins. The hypothesis of mutual interactions of both basal and metal-regulated transcription factors with the same metal-regulatory regions is discussed.

Topics: Animals; Base Sequence; Binding Sites; Cell Nucleus; Chelating Agents; DNA-Binding Proteins; Edetic Acid; Ethylenediamines; Gene Expression Regulation; Glutathione Transferase; L Cells; Metallothionein; Metals; Mice; Oligodeoxyribonucleotides; Phenanthrolines; Podophyllin; Podophyllotoxin; Promoter Regions, Genetic; Recombinant Fusion Proteins; Substrate Specificity; Transcription Factor MTF-1; Transcription Factors; Transfection