Fig. 3. The Nodal signal is not relayed indirectly between the node and LPM in mouse. (A) Schematic representation of a Cryptic transgene. An LPE isolated from a Cryptic genomic fragment was linked to the hsp68 promoter, Cryptic cDNA, an IRES, lacZ and a polyadenylation signal. Black and white boxes represent the open reading frame and untranslated regions, respectively, of Cryptic, with the arrow indicating the direction of transcription. (B,C) An E8.2 embryo harboring the transgene (Tg⁺) shows ß-galactosidase activity specifically in the LPM. (D,E) In situ hybridization for Cryptic mRNA in wild-type (D) or Cryptic^-/-; Tg⁺ (E) embryos at E8.2. Cryptic is expressed only in LPM, not in the node (black arrowhead) or floor plate, of the Cryptic^-/-; Tg⁺ embryo. (F-Q) In situ hybridization for Nodal (F-I), Lefty1 and Lefty2 (J-M) or Pitx2 (N-Q) transcripts in wild-type (F,J,N), Cryptic^-/- (G,K,O), or Cryptic^-/-; Tg⁺ (H,I,L,M,P,Q) embryos at E8.2. The expression of Nodal, Lefty2 and Pitx2 in LPM is lost in Cryptic^-/- embryos (G,K,O), but is rescued by the transgene in Cryptic^-/-; Tg⁺ embryos (H,L,P). Red arrowheads (I,M,Q) indicate ectopic gene expression in the right LPM of Cryptic^-/-; Tg⁺ embryos, which probably results from a defect in the midline barrier (asterisks in L and M). H,I,L,M are all at the five-somite stage. a, anterior; fp, floor plate; lp, LPM; p, posterior; pA, polyadenylation signal. Scale bar: 200 µm.