Functional Analysis of Agamous

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9316194 Ma The long term goal of these studies is to understand the molecular mechanisms controlling flower development in Arabidopsis. Three specific aims are identified: (1) The PI will analyze agamous (AG) function using inducible promoter fusions. The focus will be on already generated transgenic plants that carry fusions of AG with a heatshock promoter. The PI will perform temperature shift experiments at different times and study the effects of such treatment. (2) The PI will identify and characterize AG binding sites. He has characterized in vitro DNA binding with wild-type AG proteins. He will use the DNA binding assays to characterize several AG truncation mutant proteins which he has expressed in E. coli. He will also test one point mutant that causes a very interesting ag-like phenotype in transgenic plants. The PI also plans to test DNA binding in yeast and will use this system to analyze different AG target sites, including those from the AGL1 gene. (3) The PI will test possible AG target genes, using in vitro and in vivo assays to study AG binding to potential AG-binding sites in the 5' AGL1 untranscribed region and in the large AGL1 intron. Sites identified in vitro will be further analyzed in yeast and in transgenic plants using reporter gene constructs. These studies will further our understanding of AG function during flower development, particularly its role as a transcriptional regulator of genes in a regulatory hierarchy. ***