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Fig. S9. Ectopic otic vesicle formation by activation of Wnt signaling is reduced when sp8 expression is down-regulated. Ectopic otic vesicle formation was evaluated by oc90 expression. Embryos incubated with 6-bromoindirubin-3′-oxime (BIO) from stage 13/14 to stage 28 were subject to in situ hybridization analysis. Whereas control embryos showed normal oc90 expression (A), BIO-treated embryos showed increased or ectopic oc90 expression (B–D). To address whether Wnt-induced ectopic otic vesicle formation requires sp8, embryos receiving sp8 ATGMO or sp8 TALEN injection at the two-cell stage were incubated with (E) or without (F) BIO from stage 13/14 to stage 28. Interestingly, BIO-induced ectopic otic vesicles were not observed in sp8 MO or TALEN injected embryos (E), indicating sp8 is required for the Wnt-mediated ectopic otic vesicle formation. Black arrows indicate ectopic oc90 expression. In addition, we observed that oc90 expression was retained on the dkk1-injected side (H) in comparision to the uninjected side (G), suggesting that the absence of sp8 expression in dkk1- injected embryos is not due to the loss of otic vesicle tissue. Image published in: Chung HA et al. (2014) Copyright © 2014. Image reproduced with permission of the publisher and the copyright holder. This is an Open Access article distributed under the terms of the Creative Commons Attribution License. Larger Image Printer Friendly View

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