IDENTIFICATION OF OPTIMIZED TARGET SEQUENCES FOR THE GLI3 ZINC-FINGERPROTEIN

GLI3 represents an important control gene for development and differen tiation of several body structures. Reduction in gene dosage already l eads to a severe perturbation, especially of limb morphogenesis. The g ene encodes a zinc finger protein that likely functions as a transcrip tional modulator. Because the five zinc fingers should be capable of r ecognizing an extended stretch of genomic DNA, we sought to identify s equences bound by GLI3 that may facilitate the search for target genes acting downstream of GLI3. Starting from the nonamer DNA binding sequ ence of the highly related GLI protein, we employed an oligonucleotide selection protocol to determine an optimized binding sequence for the GLI3 protein. The resulting sequence bound by the GLI3 zinc fingers c onsists of 16 nucleotides and shows a high degree of similarity to seq uences bound by the GLI and tra1 proteins. Comparison with protein-DNA interactions in the known crystal structure of the GLI-DNA complex su ggests relevant interactions of additional amino acids of GLI3 with it s target site. The newly identified GLI3 target sequence should prove very useful for both the structural analysis of the protein-DNA comple x and the search for genes whose expression is subject to regulation b y the GLI3 gene product.