Fig. 4. Misexpression of CBF1 repressing form (CBF1-eve) alters the expression of SOHo1, GH6, EphA3, CBF2 and ephrin A5, but not ephrin A2. (A) Schematic representation of the wild-type and chimeric CBF1. The top drawing represents the wild-type CBF1 protein with myc tag. The repressing form of CBF1 (CBF1-eve) was constructed by fusion of the CBF1 DNA-binding domain (DNA BD) and even-skipped repression domain (eve RD). (B) Nuclear localization of CBF1 proteins. Chick embryonic fibroblasts were transfected with retrovirus vectors for myc-tagged CBF1 (a), or CBF1-eve (b). Nuclear localization of the expressed proteins was visualized by immunofluorescence using anti-myc primary antibody (red). Transfected cells were detected by expression of the viral gag protein from RCAS vector using an anti-gag antibody (green). Scale bar: 20 µm. (C) Whole-mount in situ hybridization of E3 (stage 18-20) chick embryos transfected with CBF1-eve/RCAS using antisense probes for SOHo1 (a), GH6 (b), EphA3 (c) or CBF2 (d). Insets show the normal expression of SOHo1, GH6, EphA3 or CBF2, in the control eyes. Arrowheads indicate the border of endogenous expression areas on the control side. (D) Horizontal section in situ hybridization of E8 retina transfected with CBF1-eve/RCAS using antisense probes for EphA3 (a,b), ephrin A5 (c,d) or ephrin A2 (e,f). The temporal regions of the untransfected retinae (control) are shown in the left panels (a,c,e), and those of the contralateral transfected temporal retinae of the same embryos are shown in the right panels (b,d,f). Scale bar: 100 µm.

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