Fig. 2. Gbx2 is required for repression of Otx2 by exogenous RA. (A-D) Expression of Gbx2 in wild-type control (A,B) and RA-treated embryos (C,D). (E-J) Expression of Otx2 in a wild-type control (E,F) and RA-treated wild-type embryo (G,H) and RA-treated Gbx2^–/– embryo (I,J). (C,G,I) Embryos 4 hours after RA treatment. (D,H,J) Embryos 24 hours after RA treatment. Note that by 4 hours, Gbx2 is already induced anteriorly by RA (C), whereas Otx2 is only significantly repressed by 24 hours and restricted to the most anterior tip (arrowhead) of the embryo (G,H). The repression of Otx2 by RA is inhibited in Gbx2^–/– embryos (I) and 24 hours after RA treatment Otx2 is expanded posteriorly to the presumptive r3/4 border (arrow) (J), similar to that in untreated Gbx2^–/– embryos at E8.5 (inset in J). Anterior is towards the left.