XCIRP is regulated by XTcf-3, but not by XTcf-4. (A) RT-PCR on sibling embryos (emb), animal cap explants of naive animal caps (cont), neuralized animal caps (tBr, cont) and neuralized animal caps coinjected with truncated BMP receptor (tBr) and the indicated morpholino (Tcf3Mo: XTcf-3 specific antisense morpholino oligonucleotide, Tcf4Mo: XTcf-4 specific antisense morpholino oligonucleotide) revealed that following XTcf-3 depletion, the expression of XCIRP and XCIRP2, as well as the cement gland specific marker gene XAG is robustly downregulated. The expression of HMGN1, HMGN2 and HMGX is not regulated in a Tcf-subtype specific manner. ODC shows the amplification of the house keeping gene ornithine decarboxylase, -RT is the control amplification of not reverse transcribed RNA. (B) RT-PCR on sibling embryos (emb), animal cap explants of naive animal caps (cont) and neuralized animal caps (tBr) demonstrate that the pan-neural marker gene NCAM is induced by injection of 100 pg mRNA encoding for truncated BMP receptor (tBr). H4 shows the amplification of the house keeping gene histone 4, -RT is the control amplification of not reverse transcribed RNA. (C) Sequence alignment of the Lef/Tcf specific antisense morpholino oligonucleotides together with the corresponding mRNAs, start codons of the Lef/Tcfs are indicated in bold. Tcf4Mo: XTcf-4 specific antisense morpholino oligonucleotides, Tcf3Mo: XTcf-3 specific antisense morpholino oligonucleotides, LefMo1 and LefMo2: XLef-1 specific antisense morpholino oligonucleotidess. (D-F) Specificity of the Lef/Tcf morpholinos towards their target mRNA, (D) reduction of in vitro translated S-35 labelled XLef-1 protein in the presence of the indicated morpholinos, The asterisk marks an unspecific band. (E) reduction of injected C-terminally myc-tagged XTcf-4 by coinjection of ten picomoles Tcf4Mo, but not two picomoles Tcf3Mo. The asterisks indicate unspecific staining. Gemin staining was used as loading control. (F) reduction of injected C-terminally myc-tagged XTcf-3 by coinjection of two picomoles Tcf3Mo, but not ten picomoles Tcf4Mo. Gemin staining was used as loading control.