Abstract
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.
Highlights
In addition to the well characterized modulation of AP-1 function by post-translation modifications such as phosphorylation and oxidation-reduction [16, 17], a great deal of flexibility in AP-1 function arises from the observation that AP-1 dimers can form multicomponent complexes with other unrelated transcription factors such as NF-AT, octamer, glucocorticoid receptor, and NF-B, to effect site-specific up- or down-regulation of function (18 –21)
In this current work we describe the identification, partial purification, and properties of a second active component of this 0.8 M NaI fraction as a multimeric DNA-binding protein composed of 7–17-kDa polypeptides
This factor, termed AP-1-associated factor 1 (AF-1), enhances JunD1⁄7AP-1 DNA binding by forming a tri-molecular complex with AP-1 dimers and DNA that binds with a 100-fold higher affinity than JunD alone
Summary
Vol 271, No 47, Issue of November 22, pp. 30089 –30095, 1996 Printed in U.S.A. Modulation of JunD1⁄7AP-1 DNA Binding Activity by AP-1-associated Factor 1 (AF-1)*. The AP-1 transcription factors are a ubiquitous class of gene regulatory factors that have a leading role in controlling widely diverse cellular processes [2, 3] This class of basic leucine zipper (bZip) DNAbinding proteins binds to sequences related to the pseudopalindromic AP-1 consensus site (5Ј-TGA C/G TCA-3Ј), found in numerous genes responsive to the tumor-promoting phorbol esters like 12-O-tetradecanoylphorbol-13-acetate (TPA) [4, 5]. In this current work we describe the identification, partial purification, and properties of a second active component of this 0.8 M NaI fraction as a multimeric DNA-binding protein composed of 7–17-kDa polypeptides This factor, termed AP-1-associated factor 1 (AF-1), enhances JunD1⁄7AP-1 DNA binding by forming a tri-molecular complex with AP-1 dimers and DNA that binds with a 100-fold higher affinity than JunD alone. Inducible AF-11⁄7Jun complexes can be demonstrated in nuclear extracts from mitogen-stimulated T-cells that are partially inhibited in the presence of cyclosporine A
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
