Modulation of JunD.AP-1 DNA binding activity by AP-1-associated factor 1 (AF-1).
AP-1-associated factor 1 (AF-1), is a novel protein complex that dramatically enhances the assembly of JunD-containing dimers onto AP-1 consensus sites. We describe the partial purification of AF-1 from nuclear extracts of the T-cell line MLA 144 by ionic, hydrophobic and gel filtration chromatography. AF-1 is a DNA-binding protein composed of low molecular mass polypeptides of 7-17 kDa that exists in solution as a 34-kDa complex. JunD interactions with DNA are accelerated in the presence of AF-1 through the formation of a true tri-molecular complex with JunD dimers and DNA that assembles much more rapidly on DNA than JunD alone. DNA binding analysis of AF-1 interaction with JunD.AP-1 and DNA shows that AF-1 increases the DNA binding affinity of JunD for AP-1 sites over 100-fold. DNA cleavage footprint analysis of isolated AF-1.JunD DNA complexes shows that the ternary complex makes nearly twice as many contacts with DNA than JunD dimers alone. AF-1 interacts readily, but differentially with Jun homodimers and Jun.Fos heterodimers. These findings distinguish AF-1 as a significant protein-specific modulator of AP-1.JunD in T-cells.
Powers
C
C
Laboratory of Pathology, NCI, National Institutes of Health, Bethesda, Maryland 20892, USA.
Krutzsch
H
H
Gardner
K
K
D001704:Biopolymers; D002467:Cell Nucleus / Q000378:metabolism; D002846:Chromatography, Affinity; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D019169:Jurkat Cells; D009687:Nuclear Proteins / Q000378:metabolism*; D011485:Protein Binding; D016760:Proto-Oncogene Proteins c-fos / Q000378:metabolism; D013601:T-Lymphocytes / Q000187:drug effects / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D018808:Transcription Factor AP-1 / Q000302:isolation & purification / Q000378:metabolism*
D001704:Biopolymers; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D016760:Proto-Oncogene Proteins c-fos; D018808:Transcription Factor AP-1; D004247:DNA; D013755:Tetradecanoylphorbol Acetate