ISOLATION AND CHARACTERIZATION OF THE BINDING SEQUENCES FOR THE PRODUCT OF THE ARABIDOPSIS FLORAL HOMEOTIC GENE AGAMOUS

The Arabidopsis floral homeotic gene AGAMOUS (AG) is required for norm al flower development. The deduced AG protein contains a region which shares substantial sequence similarity with the DNA-binding domains of known transcription factors, SRF (human) and MCM1 (yeast). Therefore, it is likely that AG is also a DNA-binding protein regulating transcr iption of floral genes. We describe here several experiments to charac terize AG-DNA binding in vitro. We show that AG indeed binds a DNA seq uence matching the consensus of SRF targets. Further, we have selected the AG-binding sequences from a pool of random oligonucleotides, and deduced an AG-binding consensus sequence of TT(A/T)CC(A/T)(A/t)2(T/A)N NGG(-G)(A/t)2. We have demonstrated that AG binds to the consensus reg ion of three of the oligonucleotides by footprinting analysis. Finally , we have examined AG's relative binding affinity for different sequen ces, as compared to SRF, by gel mobility shift analysis. Our results i ndicate that AG is a sequence-specific DNA-binding protein, and that t he AG-binding consensus sequence is similar to those of MCM1 and SRF.