Specific DNA recognition and intersite spacing are critical for action of the bicoid morphogen.
We examined DNA site recognition by Bicoid and its importance for pattern formation in developing Drosophila embryos. Using altered DNA specificity Bicoid mutants and appropriate reporter genes, we show that Bicoid distinguishes among related DNA-binding sites in vivo by a specific contact between amino acid 9 of its recognition alpha-helix (lysine 50 of the homeodomain) and bp 7 of the site. This result is consistent with our earlier results using Saccharomyces cerevisiae but differs from that predicted by crystallographic analysis of another homeodomain-DNA interaction. Our results also demonstrate that Bicoid binds directly to those genes whose transcription it regulates and that the amino acid 9 contact is necessary for Bicoid to direct anterior pattern formation. In both Drosophila embryos and yeast cells, Bicoid requires multiple binding sites to activate transcription of target genes. We find that the distance between binding sites is critical for Bicoid activation but that, unexpectedly, this critical distance differs between Drosophila and S. cerevisiae. This result suggests that Bicoid activation in Drosophila might require an ancillary protein(s) not present in S. cerevisiae.
Hanes
S D
SD
Department of Molecular Biology, Massachusetts General Hospital, Boston.
Riddihough
G
G
Ish-Horowicz
D
D
Brent
R
R
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D004251:DNA Transposable Elements; D004268:DNA-Binding Proteins / Q000378:metabolism*; D029721:Drosophila Proteins; D004331:Drosophila melanogaster / Q000196:embryology / Q000235:genetics / Q000378:metabolism*; D004625:Embryo, Nonmammalian / Q000378:metabolism; D005260:Female; D005801:Genes, Homeobox; D018398:Homeodomain Proteins; D007301:Insect Hormones / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D008297:Male; D008961:Models, Structural; D008969:Molecular Sequence Data; D009024:Morphogenesis; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000378:metabolism; D017433:Protein Structure, Secondary; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D015534:Trans-Activators
D004251:DNA Transposable Elements; D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D018398:Homeodomain Proteins; D007301:Insect Hormones; D009838:Oligodeoxyribonucleotides; D015534:Trans-Activators; C056478:bcd protein, Drosophila