Proposal: 9630617 PI: J. N. Petitte Primordial germ cells (PGCS) are cells that give rise to eggs or sperm and segregate from somatic lineages early in embryo development. As such, germ cells are responsible for the continuation of a species and are the genetic link between generations. Recent work on the origin of PGCs in the early avian embryo indicates that definitive germ cells begin to appear prior to gastrulation with the expression of immunological markers such as Stage-Specific Embryonic Antigen-1 (SSEA-1). Similar immunological markers are also expressed by mammalian primordial germ cells. In the chick embryo, SSEA-1 first detects PGCs at the time of oviposition prior to incubation (Stage X) and at later stages marks cells in the hypoblast (Stage Xill). SSEA-1 also identifies germ cells after gastrulation during their period of migration and following settlement in the gonad. Although the origin of the avian germ line has been studied for several decades, several questions remain concerning the time of germ cell restriction and the cellular and molecular mechanisms of avian PGCs emergence. Therefore, the objectives of this study are to determined the developmental potential of SSEA-1 positive cells of the hypoblast in vivo and to analyze the function of candidate growth factors/cytokines involved in the emergence of avian PGCS. The first goal will be assessed using the classical quail/chick embryo transplant model. SSEA-1 positive and negative quail hypoblast cells will be injected into the subgerminal cavity of stage X chicken embryos and the fate of the quail cells will be traced using a quail specific antibody. The role of cyotkines/growth factors such as stem cell factor, leukemia inhibitory factor and fibroblast growth factor-2 will be analyzed using a culture system developed in our laboratory that allows the normal processes of germ cell development to occur in vitro. Does-response effects of candidate peptide growth factors on emergence and proliferation will be assessed. As confirmation of growth factor responses , attempts will be made to inhibit the in vitro and in vivo emergence of PGCs using neutralizing antibodies, inhibitors, or anti-sense oligonucleotides to active growth factors or their receptors. This study should provide significant insights towards our understanding of germ cell development in the avian embryo and vertebrate in general.
