The AP-1 transcription factors are a ubiquitously expressed family of phorbol ester inducible proteins that play critical roles in the control of several cellular responses to extracellular stimuli including cellular growth, proliferation, and differentiation. More specifically, the AP-1 family of proteins play a significant role in the lymphoid specific cytokine expression. We have identified and purified a novel junD/AP-1 containing multi-component complex with has DNA-binding properties that are dramatically different from junD purified from other sources. We have found that many of the attributes of junD DNA binding activity in T-cells is attributable to the effect of both post-translational modification and the association of other non-jun and non-fos proteins. Together these factors combine to distinguish biochemical properties of T-cell junD in its DNA-binding stability and specificity. We have identified phosphorylation as a significant modifier of junD DNA-binding activity. In addition, we have identified and partially purified a novel factor, termed AAF-1 (AP-1 associated factor 1), that increases junD DNA binding activity greater than 100 fold. AAF-1 is a DNA binding protein that binds specifically to AP-1 and AP-1 like enhancer elements. AAF-1 enhances junD DNA binding activity by forming discrete higher order ternary complexes with DNA-bound junD. The formaton of these ternary AAF-1junD-DNA complexes is protein specific since AAF-1 inhibits rather than enhances the DNA- binding activity of c-jun DNA complexes. The reduced ability of AAF-1 to enhance the DNA binding activity of bacterially expressed junD by comparison to junD isolated from T-cells either by immuno or DNA affinity chromatography, suggest that other modification of junD in T-cells make it competent to form ternary DNA-binding complexes with AAF-1. The amino acid sequence of several proteolytic fragments of partially purified AAF-1 have been deduced and indicate that it is a novel protein(s).
