Recognition of multiple patterns of DNA sites by Drosophila homeodomain protein bicoid

Our previous studies demonstrated that the Drosophila homeodomain protein, Bicoid (Bcd), binds DNA cooperatively. In this study, we determined the patterns of adjacent DNA sites required for cooperative recognition by Bcd. Our in vitro selection and biochemical experiments demonstrated that Bcd binds preferentially to both head-to-head and tail-to-tail symmetric sites that are separated by short spacing. An increase in the spacing reduces the strict requirement of symmetric patterns of adjacent sites, permitting Bcd to recognize tandem repeat sites cooperatively. Our further experiments in vivo showed that the only pair of optimally spaced symmetric Bcd sites in a hunchback (hb) enhancer element contributes the most to transcriptional activation by Bcd, demonstrating the biological importance of the binding site patterns revealed by our in vitro selection studies.