9316986 Wasserman Dr. Wasserman is studying a conserved signal transduction pathway that regulates gene expression by controlling the subcellular localization of a transcription factor. Intracellular transduction of a signal received at the surface of the Drosophila embryo promotes nuclear import of the dorsal protein and thereby establishes the dorsoventral body axis. In mammalian T cells, the homologous pathway regulates transport of the transcription factor NF-kappaB into nuclei in response to the cytokine interleukin-1. Dr. Wasserman would like to understand where each of the known proteins in the pathway resides and how these proteins interact with one another to effect signal transduction. In addition, he would like to isolate the genes for any components in the pathway that have not been identified previously. He will use antisera directed against pathway proteins to determine their distribution in whole mount embryo preparations and then examine whether that distribution is affected by signal transduction or is dependent on the presence of other proteins in the pathway. He will also map the sequences responsible for localizing each protein, so that he may assay the role of protein localization in signaling. To investigate interactions among components of the dorsoventral pathway, he will us the two- hybrid screening system in S. cerevisiae. Pairs of Drosophila proteins will be separately fused to either LexA DNA binding domain or an acidic transcriptional activating domain and assayed for their ability to promote transcription downstream of LexA operator sites through protein-protein interaction. He will also use the yeast system to isolate mutations that prevent interaction between the Drosophila proteins. Such mutations will then be expressed in the context of the Drosophila embryo to assay their effect on signal transduction. To identify additional genes that might function in the dorsoventral signaling pathway, he will screen for Drosophila prote ins that bind to known pathway components. Using the yeast two-hybrid system, he will investigate whether any of the five previously identified proteins can interact specifically with other proteins encoded by an embryonic cDNA library. Once he has identified candidate clones in this screen, he will derive data on sequence, map position, and, where possible, mutant phenotype, to determine the likelihood that the encoded proteins play a role in transducing the dorsoventral signal. ***
