Fig. 7

Fig. 7 pitx2-EnR RNA disrupts dorsoanterior development and prevents expression of many endoderm markers. (A–C) Phenotypes caused by injection of 100 pg of the EnR-pitx2 RNA into the two dorsal vegetal cells at the eight-cell stage, including microcephaly, reduction of the size of the cement gland, and mild cyclopia (A); mild cyclopia, shortening of the axis, and spinibifida (B); and severe anterior and posterior truncations and presence of a collapsed blastocoele (b) (C). Uninjected embryos are included at the tops of A–C. (D) Embryos injected radially with 50 pg of the EnR-pitx2 RNA into each vegetal cell at the eight-cell stage and cultured until stage 27. No epiboly has occurred, gastrulation was prevented, and no dorsal or anterior structures have developed. (E–L) EnR-pitx2 RNA was injected into various blastomeres at the four- or eight-cell stage and embryos were fixed for in situ hybridization at stage 10.5–11.5. (E) Xbra expression at stage 10.5 extinguished (arrow) in half the embryo following injection of 50 pg of EnR-pitx2 RNA into two vegetal cells at the eight-cell stage (compare with Fig. 2F). (F) Similar injection with lineage tracing of the injected blastomere. Note the presence of cells that do not express Xbra and do not contain the lineage label (arrow). (G) Bisected uninjected stage 11.5 embryos showing cerberus expression throughout the endoderm, concentrated on the dorsal side. (H) Expression of cerberus is greatly reduced in siblings injected with 50 pg of EnR-pitx2 RNA into the four vegetal cells at the eight-cell stage. (I) Bisected uninjected stage 10.5 embryos showing sox-17β expression throughout the endoderm. (J) Siblings injected with 50 pg of EnR-pitx2 RNA into the four vegetal cells at the eight-cell stage showing greatly reduced expression of sox-17β. (K) Bisected uninjected stage 11 embryo showing expression of sox-17β throughout the endoderm. (L) A sibling injected with 50 pg of EnR-pitx2 mRNA into two vegetal cells at the eight-cell stage in which expression of sox-17β was abolished in half of the endoderm territory.