The prostate is an exocrine gland composed of ductal networks connected to the urethral lumen. Fetal prostate development begins when buds of urogenital sinus (UGS) epithelium invade the surrounding mesenchyme and form ducts which elongate and branch to produce a complex ductal tree. During the process of ductal morphogenesis cell proliferation, mesenchymal proliferation and mesenchymal smooth muscle differentiation are intimately coordinated. We have identified the gene Sonic hedgehog (Shh) as the likely organizer of ductal development. Shh is a secreted peptide expressed by the epithelium of the UGS. Expression is upregulated in response to testosterone and localizes to the sites of ductal budding and in the tips of the growing ducts. When Shh signaling is blocked, ductal budding is arrested, mesenchymal and epithelial proliferation are diminished, and smooth muscle differentiation is inhibited. Shh exerts its effects through a transcription factor, Gli1, which activates transcription of Shh target genes that exert specific actions on growth and differentiation. The teratogenic effects of plant steroidal alkaloids such as cyclopamine are attributed to their ability to inhibit Shh signal transduction and target gene activation. Our search for Shh target genes in the UGS identified Insulin-like growth factor binding protein-6 (IGFBP6) as a gene activated by Shh in the developing prostate. IGFBP6 is a secreted peptide that binds to and inhibits the actions of Insulin-like Growth Factor II (IGF-II). We will examine the hypothesis that IGFBP6 transcription is activated by Gli1 in response to Shh signaling and that IGFBP6 regulates cell proliferation and mesenchymal smooth muscle differentiation by inhibiting the actions of IGF-II. We will characterize IGFBP6 expression in the developing CD1 mouse prostate and determine the roles of Shh and Gli1 in activating IGFBP6 expression. The specific effects of IGFBP6 on epithelial and mesenchymal cell proliferation and smooth muscle differentiation will then be examined. Finally, we will examine the impact of plant alkaloid inhibitors of Shh signal transduction on the activation of IGFBP6 and correlate inhibition of Shh signaling in vivo with changes in IGFBP6 expression and with effects on cell proliferation, smooth muscle differentiation, and ductal budding in the developing prostate. The overall aims of these experiments are to determine the contribution of a specific target gene (IGFBP6) to the actions of Shh in prostate development and to examine the potential of plant alkaloids which inhibit Shh signaling to alter IGFBP6 expression and disrupt normal prostate growth regulation.
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