Sequences flanking the hexameric G-box core CACGTG affect the specificity of protein binding.
The CACGTG G-box motif is a highly conserved DNA sequence that has been identified in the 5' upstream region of plant genes exhibiting regulation by a variety of environmental signals and physiological cues. Gel mobility shift assays using a panel of G-box oligonucleotides differing in their flanking sequences identified two types of binding activity (A and B) in a cauliflower nuclear extract. Competition gel retardation assays demonstrated that the two types of binding activity were distinct. Type A binding activity interacted with oligonucleotides designated as class I elements, whereas type B binding activity interacted strongly with class II elements and weakly with class I elements. A third class of elements, null elements, did not exhibit any detectable binding under our assay conditions. Gel retardation analysis of nonpalindromic hybrid G-box oligonucleotides indicated that hybrid elements of the same class exhibited binding affinity commensurate with the affinity of the weaker element, hybrid class I/II elements exhibited only type B binding, and hybrid class I/null and class II/null elements did not show any detectable binding activity. These binding activities can be explained by the affinity of bZip G-box binding homo- or heterodimer subunits for G-box half sites. These experiments led to a set of classification rules that can predict the binding activity of all reported plant G-box motifs containing the consensus hexameric core. Tissue- and/or development-specific expression of genes containing G-box motifs may be regulated by the affinity of G-box proteins for the different classes of G-box elements.
Williams
M E
ME
Laboratory for Plant Molecular Biology, Rockefeller University, New York, New York 10021-6399.
Foster
R
R
Chua
N H
NH
D001483:Base Sequence; D001937:Brassica / Q000235:genetics*; D002467:Cell Nucleus / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D011485:Protein Binding; D012045:Regulatory Sequences, Nucleic Acid; D012091:Repetitive Sequences, Nucleic Acid
D004268:DNA-Binding Proteins; D004247:DNA