Understanding the molecular mechanisms underlying the order and precision of compensatory liver regeneration is essential for understanding and intervention in liver carcinogenesis and toxicology as well as development of strategies for liver cell transplantation and gene therapy. Increasing evidence indicates that liver regeneration is orchestrated by activation and repression of the activity of multiple cytokines within the liver rather than external hormones. Of the polypeptide regulators endogenous to the local liver environment, the FGF family of eight ligands and their receptors (FGF-R) (four genes, 16 splice variants resulting in >100 isoforms) has large potential for transient regulation of growth and function in both parenchymal and non-parenchymal cells. This continuation project will isolate and characterize a potentially novel parenchymal cell-specific member of the FGF ligand family extracted directly from human hepatoma cells and perfused liver in vivo. Expression and molecular basis for FGF-1-specificity and specificity in signal transduction of FGF- R4 isoforms, a major FGF-R gene expressed in liver, will be determined. Expression of splice variants of the parenchymal cell-specific FGF-R2 gene, interaction with the FGF-R4 isoforms and their-role in signal transduction will be determined in resting and regenerating liver and derived cells. Ligand-, tyrosine kinase receptor- and cell type-specific natural heparan sulfate proteoglycan components of the FGF-R complex will be isolated and characterized. A potentially novel nuclear role of a specific complex of FGF-1 and a fragment of its receptor will be investigated. The temporal expression of the TGF-beta receptor and two novel serine/threonine kinase receptors during liver regeneration in parenchymal and nonparenchymal cells will be determined. FGF-toxin conjugates will be employed to probe the role of FGF ligands and receptor isotypes in regenerating liver in vivo. The project employs combined rigorous experimental approaches which include protein micropurification, immunochemistry, recombinant technologies (bacterial, baculoviral-insect cells, transfected mammalian cells) and site-directed mutagenesis to achieve the goals of the project.
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