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Fig. 1
Ongoing Wnt signaling is detectable in differentiating melanocytes. (A) Diagram of working model for melanocyte core GRN (modified from Greenhill et al., 2011). Note, two inputs from Wnt signaling on mitfa expression, one well-established role acting alongside Sox10 driving melanocyte fate specification, and hypothetical role explored here as part of a positive-feedback loop also involving Mitfa itself. (B–G) Wnt signaling in differentiating melanocytes was assessed by scoring samples of melanocytes for detection of dGFP by immunofluorescence in Tg(top:GFP) transgenic embryos from 30 to 72 hpf. GFP-positive melanocytes (arrows) are shown by immunofluorescence (B, D, F) and DIC (C, E, G) to show melanin pigment. At each stage, 30 melanocytes in each of 15 embryos were assessed for GFP expression, in the dorsal head and throughout the dorsal and lateral trunk; numbers show the percentage of melanocytes expressing GFP (mean ± SD) at corresponding stages (n = 450 per stage). (H) Schematic showing timing of BIO treatment used. (I–L) BIO treatment enhances activity of Tg(top:GFP) reporter. BIO treatment from 15 to 40 hpf dramatically enhances dGFP expression (red arrows) in the tectum of treated (K) compared with mock-treated controls (I) and also increases dGFP signal in melanocytes of Tg(top:GFP) embryos [BIO-treated (L) compared with mock-treated control (J)]. All images, lateral views using confocal microscopy. EM, embryo medium; e: eye; T: tectum. Scale bars: (B–G) 10 μm; (I–L) 100 μm.