The hLEF/TCF-1α HMG protein contains a context-dependent transcriptional activation domain that induces the TCRα enhancer in T cells

hLEF/TCF-1 alpha is a lymphoid cell-specific HMG protein that activates the distal enhancer of the gene encoding the alpha-subunit of the T-cell receptor (TCR alpha). We have shown previously that transcriptional activation by hLEF is highly dependent on the context of its binding site within the TCR alpha enhancer. Here, we demonstrate that hLEF contains a potent transcriptional activation domain that is separate from the HMG motif and is preferentially active in T cells. We find that hLEF/GAL4 fusion proteins can activate a GAL4-substituted TCR alpha enhancer up to 50-fold in T-cell lines and are as active as GAL4/VP16 in this context. Unlike GAL4/VP16, however, hLEF/GAL4 could not activate heterologous promoters bearing only GAL4 DNA-binding sites. Thus, activation by hLEF/GAL4, like that noted previously for the native hLEF activator, was strongly influenced by the context of its DNA-binding site within the TCR alpha enhancer. Inspection of enhancer mutants suggests that trans-activation by hLEF/GAL4 is especially dependent on TCF-2, a distinct T-cell-enriched protein that binds to sequences flanking the hLEF-binding site in the enhancer. Analysis of small deletion or clustered amino acid substitution mutants in the hLEF-coding sequences identified a minimal activation region between amino acids 80 and 256 that appears to be bipartite in structure. The hLEF activation domain is not notably acid or glutamine-rich but is proline-rich and includes a motif rich in tyrosine and serine residues. We conclude that sequences outside of the hLEF HMG box mediate cell- and context-specific activation of the TCR alpha enhancer and may facilitate interactions between hLEF and other T-cell-specific factors recruited to the enhancer.