MOLECULAR GENETICS OF LIVER DEVELOPMENT

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DESCRIPTION (provided by applicant): Our goals are to identify and understand the function of genes regulating liver development. We are conducting these investigations in zebrafish, a vertebrate model system well suited for embryological and genetic studies. Over the past few years, we have completed a detailed analysis of wild-type liver development in zebrafish, and conducted a large-scale forward genetic screen for genes regulating liver development using a transgenic line expressing GFP in the endoderm and endoderm-derived organs. In this genetic screen, we identified 37 recessive mutations affecting liver development. One of the most striking mutations to come out of the screen is prometheus (prf), which appears to block hepatic specification. We have positionally cloned the prt gene and found that it encodes a Wnt2b homologue. Furthermore, gene expression and transplantation analyses indicate that cells in the lateral plate mesoderm induce hepatic specification by expressing Wnt2bb/Prt. Additional data indicate that Wnt2bb/Prt regulates hepatic specification through beta-catenin. These data about the critical role of canonical Wnt signaling in hepatic specification are surprising in light of previous work in the field that has implicated Bmp and Fgf signaling in this process. We propose to continue our studies of zebrafish liver development with the following specific aims: 1) Recover and analyze the liver mutants that were identified in our screen with the goals of making them available to the community through the stock center. 2) Investigate in more detail the role of Wnt2b, Bmp and Fgf signaling in hepatocyte specification. Our analysis of the prt mutant has led to the hypothesis that Wnt2b signaling is essential for hepatocyte specification and this hypothesis will be tested further. We will also further test the role of Bmp and Fgf signaling in hepatocyte specification. 3) Investigate in detail four mutations identified in our screen that appear to affect hepatic specification: s415, s436, s818 and s853. These mutations will be analyzed in depth including detailed phenotypic analysis and isolation of the affected gene. Altogether, these studies should provide valuable information to help us direct endodermal cells to differentiate towards the hepatocyte lineage.