Method for cloning in vivo targets of the Egr-1 transcription factor.
A methodology is described that allows the in vivo trapping of transcription factors to their target regulatory elements in multiple genes simultaneously. Cross-linking using formaldehyde is the first of several steps to isolate, purify, clone and characterize multiple gene promoter DNA fragments. The example that we use indicates that the TGF beta 1 gene is a direct target induced by Egr-1 in HT1080 cells that express constitutive Egr-1, thus explaining the growth retardation that follows Egr-1 expression. The genes identified using this procedure reflect the specific activities of Egr-1 at that moment in the cell and provide a method for confirmation of genes that are the direct targets of Egr-1 action.
de Belle
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Burnham Institute, La Jolla, CA, USA. idebelle@burnham-inst.org
Mercola
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Adamson
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ED
D001665:Binding Sites; D001943:Breast Neoplasms / Q000378:metabolism; D003001:Cloning, Molecular / Q000379:methods*; D003432:Cross-Linking Reagents; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism / Q000494:pharmacology; D051766:Early Growth Response Protein 1; D005354:Fibrosarcoma; D005557:Formaldehyde; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D017874:Immediate-Early Proteins; D007163:Immunosorbent Techniques; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism / Q000494:pharmacology; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000235:genetics; D014407:Tumor Cells, Cultured
D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; D017874:Immediate-Early Proteins; D014157:Transcription Factors; D016212:Transforming Growth Factor beta; D005557:Formaldehyde; D004247:DNA; D013755:Tetradecanoylphorbol Acetate