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. Author manuscript; available in PMC: 2010 Jul 1.
Published in final edited form as: Dev Cell. 2009 Jul;17(1):49–61. doi: 10.1016/j.devcel.2009.05.011

Figure 6. At later stages of primary hair follicle development Eda-A1/Edar/NF-κB signaling is required for maintenance of Wnt/β-catenin activity and Wnt10b expression, but not for patterned β-catenin mRNA upregulation, and Wnt10b is a potential direct NF-κB target.

Figure 6

(A) TOPGAL and cond-lacZ Wnt reporter expression analyzed by X-Gal staining in control, ΔN, and dl/dl backgrounds at E15.5. (B) Whole mount X-gal staining of E15.5 β-catenin+/lacZ (βcatlacZ), β-catenin+/lacZ;dl/dl (βcatlacZ;dl/dl), and β-catenin+/lacZ;Ta/Ta (βcatlacZ;Ta/Ta) embryos as indicated. Patches of β-catenin expression appear fused and form strings in downless and tabby mutant embryos (red arrows), as also observed in embryos of these genotypes at E13.5 and E14.5 (Figure 2C). (C) Whole mount in situ hybridization of control (c.) and ΔN embryos at the time points indicated using Wnt10b probe. Arrowheads indicate focal expression. (D) In situ hybridization for Wnt10b using sagittal sections of control and dl/dl skin at E14.0 and E15.5. (E) Whole mount in situ hybridization of E13.5 (upper panels) and E14.5 (lower panels) Ta/Ta skin explants using Wnt10b probe. Explants were treated for 24 hours with recombinant Fc-Eda-A1 (+ A1), Fc-Eda A2 (+A2), TNFα (+ TNFα), or were untreated (nt). (F) Upper panel: Conserved NF-κB binding sites in the human and murine Wnt10b promoters at the positions indicated. A verified NF-κB DNA binding site in the IκBα promoter is listed for comparison. Lower panels: ChIP using E14.5 dorsal skin extracts, primers that amplify a region encompassing the two proximal NF-κB consensus sequences in murine Wnt10b, and anti-p65 or IgG control antibodies. Primers amplifying a Gapdh promoter fragment were used as a negative control.