FUNCTION OF FIBROBLAST GROWTH FACTOR GENE IN MAMMALIAN EMBRYOGENESIS

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Evidence is rapidly accumulating that basic fibroblast growth factor 9b- FGF) and related proteins play an important role in embryonic induction. In mammals, four genes (a-FGF, k-FGF, int-2 and FGF-5) have been identified that have extensive sequence similarity to b-FGF, and more are likely to exist. We are interested in studying the role of these genes in early mammalian development. Our first set of specific goals is to determine when and where each of these genes is expressed in the developing mouse embryo by in situ RNA hybridization and protein localization, and from this information to begin to build a hypothesis about what their function in development might be. However, since the stage- and tissue-specific functions of these different growth factors will depend on the stage- and tissue- specificity of their respective receptors, we also propose to study the expression of FGF-receptor(s) in the developing embryo. In order to achieve these goals, we must have available probes for each of these mouse genes. Thus far, the only mouse gene in the FGF family that has been cloned in int-2. We have employed a variety of techniques that utilize the polymerase chain reaction (PCR) to clone cDNAs for all the known members of the mouse FGF family and have begun to preliminary examination of their expression during embryonic development. We are extending these studies to isolate hitherto unknown members of the family. In collaboration with Dr. L. Williams group, we propose to use similar approaches to obtain the mouse FGF-receptor (FGF-R) genes. As these different genes become available, we will carry out a detailed study of their expression in the developing embryo. As information on the expression of these genes begins to accumulat, it should be possible to infer something about their respective functions in development, some may display patterns of expression in the developing embyo. As information on the expression of these genes begins to accumulate, it should be possible to infer something about their respective functions in development, some may display patterns of expression consistent with a role in development of the embryonic heart or lungs. We should be aided in drawing these inferences by frequent discussions with Dr. M. Kirschner and his collaborators, who are studying the role of b-FGF in mesoderm induction in the frog. Our second set of specific goals is to obtain mice carrying mutations in the specific FGF or FGF-R genes of interest. This will be achieved by transfecting mouse embryonic stem (ES) cells in vitro with altered copies of the gene of interest and selecting those cell in which the altered allele has replaced an endogenous copy of the gene by homologous recombination. Such cells would then be used to generate mouse strains heterozygous for the altered genes. These mice could then be inter-bred to produce offspring homozygous for the targeted genetic change, and the functional consequences of the genetic alterations assessed. The results of such studies should allow us to evaluate any hypotheses we have developed on the possible funcion of the FGF and FGF-R gene familes in development.