Fig. 4. The anterior primitive streak and its derivatives are absent in Smad4 mutant embryos. Whole-mount in situ hybridization of (A,C,E,G,I,K,M,O,Q) control and (B,D,F,H,J,L,N,P,R) Sox2Cre;Smad4^CA/N mutants. (A,B) Foxa2, (C,D) Gsc and (E,F) Lim1 are expressed in the anterior primitive streak (APS) in wild-type embryos but are absent in mutants. AVE expression is unaffected. (G-L) T expression between E6.5 and E8.5. (G,H) At E6.5 T expression confirms normal formation of the posterior primitive streak and the initiation of gastrulation. (I,J) By E7.5, T expression marks the fully extended primitive streak, and emerging axial mesoderm (arrow). In mutant embryos, no T-expressing axial mesoderm is observed. (K,L) Consistent with this result, mutant embryos lack midline expression of T in the notochord at E8.5. Posterior T transcripts indicate ongoing gastrulation. (M-P) At E7.5, Nodal and Shh transcripts identify the node, but are absent in mutant embryos. (Q,R) Loss of notochord is further illustrated by the lack of midline Shh expression, although some ectopic staining is observed.