Gli3-mediated repression of Hedgehog targets is required for normal mammary development.

The Hedgehog pathway is vital for the development of many epidermal appendages, but its role in mammary development has been unclear. Here, we show that although Gli2 and Gli3 are expressed during embryonic mammary development, transcriptional reporters of positive Hedgehog signaling are absent. Nevertheless, Gli3(xt/xt) embryos show aberrant early mammary marker expression and lack two pairs of mammary buds, demonstrating that Gli3 is essential for mammary bud formation and preceding patterning events. Misactivation of the Hedgehog pathway by targeted expression of the constitutive activator Gli1, from the Gli2 promoter in Gli3(xt/+) mice, also induces mammary bud loss. Moreover, loss of Gli3 expression induces Gli1 misexpression in mammary mesenchyme. These results establish that the essential function of Gli3 during embryonic mammary development is to repress Hedgehog/Gli1-inducible targets. During postnatal mammary development, Gli2 and Gli3 are expressed in stromal and myoepithelial cells, and Gli3 is also found within the lumenal epithelium. Again, transcriptional reporters of positive Hedgehog signaling are absent from these cell types, yet are expressed robustly within mammary lymphatics. Thus, positive Hedgehog signaling is absent throughout mammary development, distinguishing the mammary gland from other epidermal appendages, such as hair follicles, which require Hedgehog pathway activity.