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A T-cell-specific transcriptional enhancer lies within the J delta 3-C delta intron of the human T-cell receptor delta gene. We have previously shown that a 30-bp element, denoted delta E3, acts as the minimal TCR delta enhancer and that within delta E3, adjacent and precisely spaced binding sites for core-binding factor (CBF/PEBP2) and c-Myb are essential for transcriptional activity. These data suggested that CBF/PEBP2 and c-Myb synergize to mediate transcriptional activity but did not establish the molecular basis for synergy. In this study, we have examined in detail the binding of CBF/PEBP2 and c-Myb to delta E3. We found that CBF/PEBP2 and c-Myb could simultaneously occupy the core site and one of two overlapping Myb sites within delta E3. However, equilibrium binding and kinetic dissociation experiments suggest that the two factors bind to delta E3 independently, rather than cooperatively. This was found to be true by using isoforms of these factors present in extracts of transfected COS-7 cells, as well as the natural factors present in nuclear extracts of the Jurkat T-cell line. We further showed that CBF/PEBP2 and c-Myb provide unique transactivation functions, since the core-Myb combination cannot be substituted by dimerized core or Myb sites. We propose that spatially precise synergy between CBF/PEBP2 and c-Myb may result from the ability of the two factors to form a composite surface that makes unique and stereospecific contacts with one or more additional components of the transcriptional machinery.

c-Myb and Core-Binding Factor/PEBP2 Display Functional Synergy but Bind Independently to Adjacent Sites in the T-Cell Receptor δ Enhancer